Antibacterial optically pure benzoquinolizine carboxylic acids, processes, compositions and methods of treatment

ABSTRACT

The present invention relates to optically pure S-(−)-benzoquinolizine carboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof, substantially free of their R-(+)-isomers, to processes for preparation of the optically pure S-(−)-benzoquinolizine carboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof substantially free of their R-(+)-isomers, and to pharmaceutical compositions comprising the S(−)-benzoquinolizine carboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof.

This application claims the benefit of U.S. Provisional Application No.60/170,676 Dec. 14, 1999 and incorporates the same by reference.

FIELD OF THE INVENTION

The present invention relates to optically pure S-(−)-benzoquinolizinecarboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphsand hydrates thereof, substantially free of their R-(+)-isomers, toprocesses for preparation of the optically pure S-(−)-benzoquinolizinecarboylic acids, their derivatives, salts, pseudopolymorphs, polymorphsand hydrates thereof substantially free of their R-(+)-isomers, and topharmaceutical compositions comprising the S(+)-benzoquinolizinecarboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphsand hydrates thereof. These compounds and compositions can be used tosystemically and topically treat bacterial Gram-positive, Gram-negativeand anaerobic infections, specially resistant Gram-positive organisminfections, Gram-negative organism infections, mycobacterial infectionsand emerging nosocomial pathogen infections, while avoiding toxiceffects associated with the administration of the racemic mixture ofRS-(±)-benzoquinolizine carboxylic acid. The compounds and compositionsof this invention can also be need to treat diseases and disorderscaused by Gram-positive, Gram-negative and anaerobic bacteria, anddiseases and disorders caused by resistant Gram-positive organisms,Gram-negative organisms, mycobacteria and nosocomial pathogens.

BACKGROUND OF THE INVENTION

Bacterial resistance to antibiotics is an increasingly recurrentphenomenon. Of grave concern has been the development ofmethicillin-resistant Staphylococcus aureus (MRSA) andmethicillin-resistant Streptococcus epidermidis (MRSE) strains, whichbecause of the phenomenon of cross-resistance, are now also resistant tothe larger class of β-lactam antibiotics including the cephalosporinsand carbapenems. Of even graver concern is the development of resistancein MRSA strains against the class of antibacterial agents known asfluoroquinolones. Several reports are known of MRSA strains displayingresistance to fluoroquinolone agents such as ciprofloxacin, sparfloxacinand even the more recently introduced trovafloxacin. In addition, fortrovafloxacin and for newer introductions like grepafloxacin,moxifloxacin and gatifloxacin, a concern has been expressed about theircheckered safety records. The use of trovafloxacin has been suspended orseverely curtailed because of its association with liver side effects.Grepafloxacin was withdrawn worldwide because of severe cardiovascularside effects. The labelling on gatifloxacin and moxifloxacin warns thatthey may prolong the QTc interval on electrocardiograms in somepatients.

The last line of defense against such fluoroquinolone-resistant MRSAstrains is the class of glycopeptide antibiotics represented byvancomycin and teicoplanin. These glycopeptide antibiotics are, however,laden with several limitations. Vancomycin is encumbered with lack oforal bioavailability, nephrotoxic potential, toxic effects such asphlebitis and red-men syndrome. Moreover, the recent disturbing widespread emergence of Vancomycin resistant enterococci (VRE) followed bythe alarming reports of Vancomycin intermediate resistanceStaphylococcus aureus (VISA) strains from Japan and USA have cast ashadow over the future of glycoside antibiotics in clinical practice. Intime, there is a relatively wide-spread emergence of staphylococci,enterococci, pneumococci and streptococci, which have become resistantto currently used first-and second-line antibacterial agents such aspenicillin, oxacillin, vancomycin and erythromycin (SENTRY Programme:Antimicrobial Agents & Chemotherapy 42 1762-1770,1998).

Also, for primary skin infections such as impetigo and folliculitis, andfor secondary infections in humans such as infected dermatitis, woundsand burns, as well as to eliminate nasal carriage of MRSA in healthcareworkers and patients, a special antibiotic used topically is Mupirocin.Mupirocin has high in vitro anti-staphylococcal and anti-streptococcalactivity. There has, however, been an increase of organisms, speciallystaphylococci, developing resistance to Mupirocin. The emergence ofMupirocin-resistant Methicillin-resistant Staphylococcus aureus (MRSA)in infected patients in different countries like Canada, WesternAustralia, UK, Spain and Switzerland is described in differentreferences in the medical and scientific literature viz. J. Hosp.Infect. 39(1), 19-26 (1998); J. Hosp. Infect. 26(3),157-165 (1994);Infect Control Hosp Epidemiol 17(2), 811-813 (1996); 38^(th) AnnualICAAC Abstract C-75, 90 (1998); 38^(th) ICAAC Abstract 12-25, 507(1998).

Furthermore, Gram-positive pathogens such as Staphylococci, enterococciand Gram-negative pathogens E. coli, Klebsiella, Proteus, Serratia,Citrobacter and Pseudomonas, frequently encountered in urinary tractinfections are susceptible to the known fluoroquinolones, such asciprofloxacin, levofloxacin, ofloxacin and norfloxacin. The potency ofthese fluoroquinolones, however, markedly deteriorates under the acidicconditions likely to be encountered in urinary tract infections,rendering them inadequate.

Furthermore, multidrug-reistant (DR) mycobacterial strains have emergeddisplaying resistance to first-line antimycobacterial agents such asrifampicin, pyrazinamide and INH etc. thus severely curtailingtherapeutic options available for the management of infections due tosuch strains. Usually, the antimycobacterial drug regimen involvestreatment spread over several months, and hence the drug has to betolerated well by the patients. Among the fluoroquinolone antibiotics,sparfloxacin is reported to be highly active against mycobacteria. It isnot quite suitable, however, for long-term therapy because of itspotential to cause phototoxic side effects in humans and laboratoryanimals such as mice and guinea-pigs.

Furthermore, in the worldwide management of nosocomial infections,besides the problematic strains of staphylococci and enterococci,including MRSA, strains of Chryseobacteria have recently emerged as newmembers of nosocomial pathogens causing neonatal meningitis andpneumonia, as well as sepsis, in immuno-compromised patients beingtreated in intensive care units. Chryseobacteria are intrinsicallyresistant to β-lactam antibiotics including third-generationcephalosporins and carbapenems. These factors reduce the treatmentoptions available to the clinicians.

The highly pressing need for other agents and methods of treatment forinfections arising from such emerging resistant microorganisms,Gram-negative pathogens in acidic environments, mycobacteria andnosocomial pathogens thus assumes great significance.

Among other agents, one particular class of compounds thebenzoquinolizine carboxylic acids are of particular relevance.Nadifloxacin is an example of a benzoquinolizine carboxylic acid.Nadifloxacin is racemic[(±)-9-fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H-5H-benzo[i,j]quinolizine-2-carboxylic acid and is disclosed in JP Patent No.58,90,511 and U.S. Pat. No. 4,399,134. Nadifloxacin has an asymmetriccarbon atom at the 5-position thereof. RS-(±)-Nadifloxacin comprises twooptically active isomers. In describing an optically active compound,the prefixes R and S or D and L are used to denote the absoluteconfiguration of the molecule about its chiral centre(s). The prefixes(+) and (−) or d and l are employed to designate the sign of rotation ofplane-polarized light by the compound, with (−) or l meaning that thecompound is levorotatory. A compound prefixed with (+) or d isdextrorotatory. Compounds having a single chiral centre exist as a pairof enantiomers, which are identical except that they arenon-superimposable mirror images of one another. A one-to-one mixture ofenantiomers is often referred to as a racemic mixture. RacemicRS-(±)-Nadifloxacin derives its biological activity primarily from theS-(−)-enantiomer. The optically active S-(−)-Nadifloxacin [α]²⁰_(D)=−312.0 is obtained as disclosed in Chem. Pharm. Bull 44 (1996),page nos. 642-5 and Jpn. Kokai Tokyo Koho JP 63,192,753. The opticallyactive R-(+)-Nadifloxacin, [α]²⁰ _(D)=+312.0, is obtained as disclosedin Jpn.Kokai Tokyo Koho JP 63,192,753. Pharmaceutical compositions ofRS-(±)-Nadifloxacin are disclosed in U.S. Pat. No. 4,399,134 and U.S.Pat. No. 4,552,879. Although these cited patents disclose compositionsof RS-(±)-Nadifloxacin for oral, parenteral and topical use, the onlycommercial product containing RS-(±)-Nadifloxacin as an activeantibacterial compound is the commercial product named Acuatim®.Acuatim® is available as a cream and a lotion and incorporates racemicRS-(±)-Nadifloxacin as 1% of its composition for the topical treatmentof acne. Acuatim® has several drawbacks. It is intended only for topicaluse and is registered only for the treatment of acne caused byPropionibacterium species. One report has appeared on the in-vitroactivity of the fluoroquinolone, Nadifloxacin, against methicillinresistant isolates of Staphylococcus aureus from patients with skininfections (see Nishijima et al., Drugs 49 (Suppl.) 230-232, 1995).There is no report of RS-(±)-Nadifloxacin being approved for systemicuse against any microbial infections, whether for sensitive or resistantmicrobial strains.

S-(−)-Nadifloxacin is reported in Chem. Pharm, Bull 44 (1996) pages Nos.421-423 to be approximtely twice as active in-vitro as racemicNadifloxacin against Gram-positive and Gram-negative bacteria. There isno previous report, however, of the activity of S-(−)-Nadifloxacin inin-vivo systems against Gram-positive bacteria, Gram-negative bacteria,anaerobes, mycobacteria and emerging nosocomial pathogens.

RS-(±)-Nadifloxacin is reported to exist in two crystalline forms, oneas an anhydrate and the other as a hemihydrate (M. Kido and K Hashimoto,Chem. Pharm. Bull, 42, 872 (1994)). There is no previous report,however, of any hydrate forms of S-(−)-Nadifloxacin, although anon-hydrate form is reported (K. Hashimioto et al., Chem. Pharm. Bull.,44,642 (1996)).

There is no previous report of the utility of optically purebenzoquinolizine carboxylic acids, their derivatives, salts,pseudopolymorphs, polymorphs and hydrates thereof of the invention inpharmaceutical compositions. There is also no previous report of thesystemic or topical use of optically pure benzoquinolizine carboxylicacids, their derivatives, salts and hydrates thereof of the invention,either alone or in compositions for treatment of microbial infectionsdiseases or disorders.

Our pending PCT patent application No. PCT/IN99/00016 filed on May7,1999 describes optically pure and racemic benzoquinolizine carboxylicacids, derivatives and salts thereof for treatment of infections causedby Mupirocin-resistant bacterial strains such as Mupirocin-resistantstaphylococci, Mupirocin-resistant streptococci and otherMupirocin-resistant Gram-positive and Mupirocin-resistant Gram-negativebacteria, and for treatment of dermal diseases such as impetigo,folliculitis, infected dermatitis, wounds and burns. The subject matterof PCT application PCT/IN99/00016 is incorporated herein by reference.

SUMMARY OF THE INVENTION

The present inventors obtained optically pure isomers of Nadifloxacinand have conducted extensive studies to show that:

-   -   1. S-(−)-Nadifloxacin is found to exist not only as an anhydrate        but also as three new different hydrates designated as        S-(−)-Nadifloxacin.nH₂O, wherein n is equal to 0.2, 0.5 or 0.75.        Among these forms of the anhydrate and the three hydrates,        S-Nadifloxacin.0.2 H₂O is now specifically found to be preferred        as a stable, non-hygroscopic crystalline modification which is        distinguished by an increased stability, neither losing the        water content therein nor absorbing moisture over a wide range        of ambient relative humidity conditions. In contrast, moisture        absorption by the anhydrate varies according to its method of        preparation and the relative humidity conditions to which it is        subjected. The other two hydrates, viz. the hemihydrate,        S-(−)-Nadifloxacin.0.5H₂O and the hemisesquihydrate,        S-nadifloxacin.0.75H₂O, both revert to S-nadifloxacin.0.2H₂O        when dried in vacuo at ambient temperature conditions.    -    S-(−)-Nadifloxacin.0.2H2O has considerable advantages over the        anhydrate, the hemihydrate and the hemisesquihydrate in storage        and handling and in the preparation of medicament forms. In        particular, specially in tropical and subtropical climates,        where the ambient humidity is usually greater than 70%,        difficulty is encountered in the storage and handling of the        anhydrate. In the preparation of pharmaceutical preparations,        such as tablets, containing the anhydrate, the operations must        be carried out with attention to absorption or desorption of        water of crystallisation. More specifically, a room in which to        handle the anhydrate must be kept at low humidity and        conversely, a room in which to handle the hemihydrate and        hemisesquihydrate must be kept at low temperatures and high        humidity. Unless these conditions are provided, these compounds        or preparations containing these compounds would change in        weight, and thus would not serve for practical purposes and        would lose their commercial value. By using a stable        non-hygroscopic, free-flowing active compound, as is provided by        S-(−)-Nadifloxacin 0.2 H₂O, a satisfactory dosing consistency        and accuracy is achieved during the preparation of medicaments,        which increases safety and therefore minimizes the risk to the        patient.    -   2. Crystalline salts of S-(−)-Nadifloxacin, especially sodium,        potassium, and arginine salts, and hydrates thereof have been        identified with increased aqueous solubility over        S-(−)-Nadifloxacin, and consequently with superior properties        for use in the preparation of parenteral formulations, and with        advantages of improved oral bioavailability in solid oral dosage        forms.    -   3. Derivatives of S-(−)-Nadifloxacin are identified at the sites        of the 2-COOH function and the 4′-hydroxy moiety of the        9-(4′-hydroxypiperidino) group respectively, and salts and        hydrates thereof.    -   4. Processes are described to obtain the optically pure        benzoquinolizine carboxylic acids of the invention, the        derivatives, salts, pseudopolymorphs, polymorphs and hydrates        thereof mentioned under items 1-3 above and as described in        detail below.    -   5. RS-(±)-Nadifloxacin, S-(−)-Nadifloxacin and optically pure        benzoquinolizine carboxylic acids, the derivatives, salts,        pseudopolymorphs, polymorphs and hydrates thereof have high        activity against Mupirocin-resistant microbial strains such as        Mupirocin-resistant staphylococci, Methicillin-resistant        staphylococcus aureus and Quinolone-resistant Staphylococcus        aureus, coagulase negative staphylococci, such as        Methicillin-resistant Staphylococcus epidermidis (MRSE),        enterococci, betahemolytic streptococci and viridans group of        streptococci.    -   6. RS-(±)-Nadifloxacin, S-(−)-Nadifloxacin and optically pure        benzoquinolizine carboxylic acids, the derivatives, salts,        pseudopolymorphs, polymorphs and hydrates thereof have activity        against mycobacteria and newly emerging nosocomial pathogens        such as Chryseobacterium meningosepticum.    -   7. S-(−)-Nadifloxacin and optically pure benzoquinolizine        carboxylic acids, the derivatives, salts, pseudopolymorphs,        polymorphs and hydrates thereof have 2-4 times higher        antimicrobial activity than racemic-Nadifloxacin against        Mupirocin-resistant staphylococci, Methicillin-resistant        Staphylococcus aureus (MRSA), Quinolone-resistant Staphylococcus        aureus, coagulase negative staphylococci, such as        Methicillin-resistant Staphylococcus epidermidis (MRSE),        enterococci, betahemolytic streptococci and viridans group of        streptococci, mycobacteria and newly emerging nosocomial        pathogens such as Chryseobacterium meningosepticum.    -   8. S-(−)-Nadifloxacin and optically pure benzoquinolizine        carboxylic acids, the derivatives, salts, pseudopolymorphs,        polymorphs and hydrates thereof are not only bacteriostatic but        also bactericidal towards Mupirocin-resistant staphylococci,        Methicillin-resistant Staphylococcus aureus (MRSA),        Quinolone-resistant Staphylococcus aureus, coagulase negative        staphylococci, such as Methicillin-resistant Staphylococcus        epidermidis (MRSE), enterococci at concentrations 2-4 times        lower than that of RS-(±)-Nadifloxacin.    -   9. S-(−)-Nadifloxacin and optically pure benzoquinolizine        carboxylic acids, the derivatives, salts, pseudopolymorphs,        polymorphs and hydrates thereof have 2-4 times higher        antimicrobial activity against Gram-positive pathogens such as        staphylococci and enterococci and Gram-negative pathogens such        as E. coli, Klebsiella, Proteus, Serratia and Citrobacter in the        acidic environments encountered in infection such as urinary        tract infections.    -   10. S-(−)-Nadifloxacin, its derivatives, salts,        pseudopolymorphs, polymorphs and hydrates thereof have high        potency against efflux pump-bearing Staphylococcus strains and        are thus of unique value in treating infections caused by        antibiotic-resistant microorganisms for which the resistance        mechanism is due to the presence of efflux pumps.    -   11. S-(−)-Nadifloxacin, its derivatives, salts,        pseudopolymorphs, polymorphs and hydrates thereof have high        propensity to display resistance to resistance development,        which has been shown in studies involving sequential        transfers/passages of a S. aureus strain through respective drug        containing media.    -   12. The acute intravenous toxicity of S-(−)-Nadifloxacin and its        arginine salt is significantly lower than RS-(±)-Nadifloxacin        (Biological Example 3).    -   13. S-(−)-Nadifloxacin, its derivatives, salts and hydrates        thereof have a favourable toxicity profile in comparison with        other fluoroquinolone drugs in clinical use in respect of        cytotoxic effect on various cell lines (Biological Example 4),        phototoxicity (vide infra) and cardiotoxicity.    -   14. The oral bioavailability of S-(−)-Nadifloxacin is 2-times        higher than that of RS-(±)-Nadifloxacin (Biological Example 5).

Through their extensive studies, the present inventors have shown forthe first time a novel expanded set of clinically desired antimicrobialattributes of efficacy and safety of S-(−)-Nadifloxacin, which have beennot reported in the literature since the first disclosure ofS-(−)-Nadifloxacin in JP 63, 192, 753 about twelve years ago and ofRS-(±)-Nadifloxacin in JP 58,90,511 about twenty years ago. Newhydrates, salts, derivatives, pseudopolymorphs, polymorphs andcompositions of S-(−)-Nadifloxacin have also been identified by thepresent inventors, which have in addition to their biological propertiesmentioned above, newer physico-chemical properties, thus permittingtheir utility in a clinical and commercial exploitation in newercompositions for newer diseases and newer methods of systemic andtopical treatment that were hitherto not possible.

It is, thus, an object of the present invention to provideS-(−)-optically pure benzoquinolizine carboxylic acids, theirderivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof,of the formula I, substantially free of their R-(+)-isomers.

It is another object of the present invention to provide a process orprocesses for preparing the novel optically pure S-(−)-optically purebenzoquinolizine carboxylic acids, their derivatives, salts,pseudopolymorphs, polymorphs and hydrates thereof, of the formula I.

A further object is to provide pharmaceutical compositions comprisingoptically pure S-(−)-benzoquinolizine carboxylic acids, the derivatives,pseudopolymorphs, hydrates and salts thereof as potent antibacterialagents for treating systemic and topical bacterial infections,especially infections caused by resistant Gram-positive, sensitive andresistant Gram-negative organisms, mycobacterial infections andnosocomial pathogen infections while avoiding the toxic effectsassociated with the administration of their R-(+)-isomers.

Another object of this inventions relates to a method of treatment ofinfections caused by Mupirocin-resistant bacterial strains such asMupirocin-resistant staphylococci, Mupirocin-resistant streptococci andother Mupirocin-resistant Gram-positive and Mupirocin-resistantGram-negative bacteria, and of dermal diseases and disorders such asimpetigo, folliculitis, infected dermatitis, wounds and burns. Treatmentcomprises oral, parenteral, administration and/or topical application ofan effective amount of a composition of S-(−)-Nadifloxacin or opticallypure benzoquinolizine carboxylic acids, their derivatives, salts,pseudopolymorphs, polymorphs and hydrates thereof of formula I, or of acomposition of RS-(+)-Nadifloxacin and pharmaceutically acceptable saltsthereof.

A further object of the invention includes methods for treating theinfections in humans and animals caused by Gram-positive, Gram-negativeand anaerobic bacteria, resistant Gram-positive organism such asMethicillin-resistant Staphylococcus aureus (MRSA), Quinolone-resistantStaphylococcus aureus, coagulase negative staphylococci, such asMethicillin-resistant Staphylococcus epidermidis (MRSE), enterococci,betahemolytic streptococci and viridans group of streptococci,mycobacteria and newly emerging nosocomial pathogens such asChryseobacterium meningosepticum, and Gram-negative pathogens such as E.coli, Klebsiella, Proteus, Serratia Citrobacter and Pseudomonas, whileavoiding the toxic effects that are associated with the racemic mixtureof Nadifloxacin by administering systemically or topicallyS-(−)-Nadifloxacin or optically pure S-(−)-benzoquinolizine carboxylicacids, their derivatives, salts, pseudopolymorphs, polymorphs andhydrates thereof to the affected human or animal.

BRIEF DESCRIPTION OF THE DRAWINGS

For a thorough understanding of the nature and objects of the invention,reference should be made to the following detailed description taken inconnection with the accompanying drawings in which

1. FIG. 1 represents the moisture content ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid anhydrate at a relative humidity of 20% to 95%.

2. FIG. 2 represents the equilibrium moisture content ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.2 hydrate at a relative humidity of20% to 95%.

3. FIG. 3 represents the X-ray diffraction pattern ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid anhydrate.

4. FIG. 4 represents the X-ray diffraction pattern ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.2 hydrate.

5. FIG. 5 represents the X-ray diffraction pattern ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.5 hydrate.

6. FIG. 6 represents the X-ray diffraction pattern ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.75 hydrate.

7. FIG. 7 represents the results of Differential Scanning Calorimetry onS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H -benzo [i,j]quinolizine-2-carboxylic acid anhydrate

8. FIG. 8 represents the results of Differential Scanning Calorimetry onS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.2 hydrate.

9. FIG. 9 represents the results of Differential Scanning Calorimetry onS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.5 hydrate.

10. FIG. 10 represents the results of Differential Scanning CalorimetryonS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.75 hydrate.

DETAILED DESCRIPTION OF THE INVENTION

This invention relates to S-(−)-optically pure benzoquinolizinecarboxylic acids, of formula

and their pharmaceutically acceptable salts, derivatives,pseudopolymorphs, polymorphs and hydrates, substantially free of theirR-(+)-isomers;wherein

X is OR₁, wherein R₁ is hydrogen, a pharmaceutically acceptable cation,such as those of alkali metals such as lithium, sodium, potassium;alkaline earth metals such as magnesium or calcium, aluminum, ammoniumor substituted ammonium salts; choline or organic amines such asdiethanolamine, guanidine or heterocyclic amines such as piperidine,hydroxyethylpyrrolidine, hydroxyethylpiperidine, morpholine, piperazine,N-methyl piperazine and the like or basic amino acids such as opticallypure and racemic isomers of arginine, lysine, histidine, tryptophan andthe like;

or R₁ is C₁-C₆ alkyl, such as straight chain or branched chain aliphaticresidues such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl,pentyl, hexyl and their branched chain isomers;

or R₁ is —(CH₂)n-CHR₄—OOCR₅, wherein R₄═H, or CH₃; n=0-3 and R₅═C₂H₅ orC(CH₃)₃, R₁ is a group such as acetoxymethyl, pivaloyloxymethyl,pivaloyloxyethyl group;

or R¹ is

wherein A=CH or N, and when A=CH, Z=NH or NCH₃, and when A=N, Z=CH, O,N, S, or NCH₃; p=0-2; q=0-2, wherein R¹ is a group such asN-methylpiperidin-4-yl, pyrrolidin-2-yl-ethyl, piperidin-2-yl-ethyl, ormorpholin-2-yl-ethyl;

or X is NHR₂, wherein R₂ is hydrogen or NHR₂ is the residue of one ofthe 20 naturally occurring amino acids: alanine, arginine, asparagine,aspartic acid, cysteine, glutamine, glutamic acid, glycine, histidine,isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine,threonine, trypotophan, tyrosine or valine or the optically activeisomers thereof, or the racemic mixtures thereof,

R₃ is hydrogen, C₁-C₆ alkyl (C₁-C₆ alkyl is defined as above), glycosyl,aralkyl such as benzyl, C₁-C₆ alkanoyl such as acetyl, propionyl,pivaloyl; or aminoalkanoyl. The amino alkanoyl group may be an aminoacid residue derived from one of the 20 naturally occurring amino acidsor the optically active isomers thereof, or the racemic mixturesthereof. The amino acid residue is derived from alanine, arginine,asparagine, aspartic acid, cysteine, glutamine, glutamic acid, glycine,histidine, isoleucine, leucine, lysine, methoinine, phenylalanine,proline, serine, threonine, tryptophan, tyrosine or valine. R₃ may alsobe C₆H₁₁O₆, PO₃H₂ or SO₃H thus giving respectively, esters with gluconicacid, phosphoric acid derivatives, or sulphuric acid derivatives of thecompounds.

y denotes an integer from 0 to 3 and any fractional numbers thereindepending on the moles of acid added to the basic molecule;

z denotes moles of water, for example 0, 0.2, 0.25, 0.5, 0.75, 1, 1.25,1.5, 2, 3, etc; and

HB represents an acid as defined below for acid addition salts.

Pharmaceutically acceptable salts are those salts already included bydefinition of the symbol X in Formula I. In addition, in view of thebasic character of the compounds of Formula I and of the basic aminoacids used in the preparation of derivatives it is possible to make acidaddition salts. Also, because of the acidic character introduced in thederivatives of Formula I, it is also possible to make basic or alkaliaddition salts of the compounds of Formula I. Preferred acid additionsalts are those of hydrochloride, hydrobromide, hydroiodide, sulphate,phosphate and salts of organic acids such as acetate, lactate,succinate, oxalate, maleate, fumarate, malate, tartrate, citrate,ascorbate, cinnamate, gluconate, benzoate, methane sulfonate andp-toluene sulfonate. Preferred alkali addition salts are lithium,sodium, and potassium salts, and alkaline earth salts are magnesium, andcalcium salts.

Specific compounds of the invention are:

-   1.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid.-   2.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid 0.2 H₂O.-   3.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid 0.5 H₂O.-   4.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid 0.75 H₂O.-   5.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid sodium salt.-   6.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, sodium salt monohydrate.-   7.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, potassium salt monohydrate.-   8.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, arginine salt.-   9.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, arginine salt 0.25 H₂O.-   10.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, arginine salt .0.75 H₂O.-   11.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, lysine salt monohydrate.-   12.    S-(−-)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, histidine salt 0.2 H₂O.-   13.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, hydroxyethyl pyrrolidine salt.-   14.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, diethanolamine salt.-   15.    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, choline salt and its hydrates.-   16. Carboxymethyl    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate    sodium salt.-   17. Acetoxymethyl    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate.-   18. Propionoxymethyl    S-(−)-9-fluoro-6,7-dihydro-8(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate.-   19. Pivaloyloxymethyl    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate.-   20. Pivaloyloxyethyl    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate.-   21. N-methylpiperidin-4-yl S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxy    piperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate.-   22. Pyrrolidin-2-yl-ethyl S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxy    piperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate.-   23. Piperidin-2-yl-ethyl    S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate.-   24. Morpholin-2-yl-ethyl S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxy    piperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate.-   25.    9-fluoro-8(4-hydroxypiperidin-1-yl)-(5S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-12(S)-amino-1,5-pentanedioic    acid] carboxamide, disodium salt.-   26.    9-fluoro-8-{4-hydroxypiperidin-1-yl}-5(S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-[2(S)-amino-3-imidazolylpropionic    acid] carboxamide hydrochloride.-   27.    S-(−)-9-fluoro-6,7-dihydro-8-(4-methoxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   28.    S-(−)-9-fluoro-6,7-dihydro-8-(4-[(β-D-tetraacetylglucopyranosyl)oxy]-piperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   29.    S-(−)-9-fluoro-6,7-dihydro-8-(4-[(β-D-glucopyranosyl)oxy]-piperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   30.    S-(−)-9-fluoro-6,7-dihydro-8-(4-acetoxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   31.    S-(−)-9-fluoro-6,7-dihydro-8-(4-pivaloyloxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   32.    S-(−)-9-fluoro-8-[4-(phosphonoxy)-1-piperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid.-   33.    8-{4-[2(S)-Amino-propionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   34.    8-{4-[2(S)-Amino-propionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, hydrochloride.-   35.    8-{4-[2(R)-Amino-propionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   36.    8-{4-[2(R)-Amino-propionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,    5H-benzo[i,j]quinolizine-2-carboxylic acid, acetate.-   37.    8-{4-[2(RS)-Amino-propionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   38.    8-{4-[(2S)-amino-propionyl-2S)-aminopropionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid hydrochloride.-   39. 8-{4 [(2R)-Amino-propionoyl-(2R)-aminopropionyloxy]    piperidin-1-yl}-9-fluoro-(5S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   40. 8-{4[(2R)-Amino-propionoyl-(2R)-aminopropionyloxy]    piperidin-1-yl}-9-fluoro-(5S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid hydrochloride.-   41.    8-{41(2R)-Amino-propionoyl-(2R)-aminopropionyloxy]piperidin-1-yl}-9-fluoro-(5S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid acetate.-   42.    8-{4[(2S)-Methylamino-propionyloxy]piperidin-1-yl}-9-fluoro-(5S)-methyl-6,7    dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid.-   43. 8-{4[(2S)-Methylamino-propionyloxy]    piperidin-1-yl}-9-fluoro-45S)-methyl-6,7    dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid and    hydrochloride.-   44. 8-{4[2(S)-    amino-3-carboxypropionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid hydrochloride.-   45.    8-{4[2(S)-amino-3-phenylpropionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid hydrochloride.-   46.    8-{4[2(R)-amino-3-phenylpropionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid hydrochloride.-   47.    8-{4[2(R)-amino-3-phenylpropionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid acetate.-   48. 8-{4[(2S)-    Amino-3-methylbutanoyloxy]piperidin-1-yl}-9-fluoro5S)-methyl-6,7    dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid.-   49.    8-{4[(2S)-Amino-3-methylbutanoyloxy]piperidin-1-yl}-9-fluoro45S)-methyl-6,7    dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid and    hydrochloride.-   50.    8-{4-[2(S)-Amino-4-methylpentanoyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   51.    8-{4-[2(S)-Amino-4-methylpentanoyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, hydrochloride.-   52.    8-{4[(2R)-Amino-4-methylpentanoyloxy]piperidin-1-yl}-9-fluoro-(5S)-methyl-6,    7 dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid.-   53.    8-{4[(2R)-Amino4-methylpentanoyloxy]piperidin-1-yl}-9-fluoro(5S)-methyl-6,    7 dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid and    hydrochloride.-   54.    8-{4-[2(S),6-Diaminohexanoyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   55.    8-{4-[2(S),6-Diaminohexanoyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, dihydrochloride.-   56. 8-{4    [(2S)-Amino-5-nitroguanidino-butanoyloxy]piperidin-1-yl}-9-fluoro-)5S)-methyl-6,7    dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid.-   57.    8-{4[(2S)-Amino-5-nitroguanidino-butanoyloxy]piperidin-1-yl}-9-fluoro-(5S)-methyl-6,7    dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid and    hydrochloride.-   58. 8-{4 [(2S)-Amino-5-guanidino-butanoyloxy]    piperidin-1-yl}-9-fluoro-45S)-methyl-6,7    dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid.-   59. 8-{4[(2S)-Amino-5-guanidino-butanoyloxy]    piperidin-1-yl}-9-fluoro-(5S)-methyl-6,7    dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid and    hydrochloride.-   60.    8-{4[(2S)-Amino-5-nitroguanidino-butanoyl-(2S)-amino-5-nitroguanidino-butanoyloxy]piperidin-1-yl}-9-fluoro45S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   61.    8-{4[(2S)-Amino-5-nitroguanidino-butanoyl-(2S)-amino-5-nitroguanidino-butanoyloxy]piperidin-1-yl}-9-fluoro-(5S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid and hydrochloride.-   62.    8-{4[(2S)-Amino-5-guanidino-butanoyl-(2S)-amino-5-guanidino-butanoyloxy]piperidin-1-yl}-9-fluoro-(5S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid.-   63.    8-{4[(2S)-Amino-5-guanidino-butanoyl-(2S)-amino-5-guanidino-butanoyloxy]piperidin-1-yl}-9-fluoro5S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid and hydrochloride.

The hydrates of all the above compounds are also compounds of theinvention. Hydrate may be represented by the term hydrate or H₂O.

One embodiment of the invention are the salts and hydrates ofS-(−)-Nadifloxacin. Another embodiment of the invention are thederivatives and salts of the S-(−)-benzoquinolizine carboxylic acids ofthe invention which are essentially prodrugs of compounds of the formulaI having free carboxylic acid groups or hydroxy groups. Prodrugs areunderstood to be esters of the free carboxylic acid group, or amides ofthe free carboxylic acid group with ammonia, organic amines or the aminogroup of an amino acid residue, or a polypeptide chain of two or more,such as up to four, amino acids residues which are covalently joinedthrough peptide bonds. Prodrugs are also understood to be ethers of thefree 4-OH group of the piperidinyl moiety or esters of the free4-OH-group of the piperidinyl moiety with a carboxylic acid residue asdefined for formula I above or with the carboxylic acid group of anorganic acid, organic dibasic acid or an amino acid residue, or apolypeptide chain of two or more, such as up to four, amino acidresidues which are covalently joined through peptide bonds. The aminoacid residues of use include the 20 naturally occurring amino acidsalanine, arginine, asparagine, aspartic acid, cysteine, glutamine,glutamic acid, glycine, histidine, isoleucine, leucine, lysine,methionine, phenylalanine, proline, serine , threonine, tryptophan,tyrosine or valine. Preferred amino acid residues are those with abasic-polar group such as Nitro-Arg, Arg, Lys, His, and those with apolar group such as Ala, Val, Nval, Leu, Met, Gly, Pro, Phe. Prodrugs atthe free 4-OH group may also be phosphoric acid esters and sulfonic acidesters.

Particularly, the preferred compounds of the present invention are:

-   S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid-   S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid 0.2 hydrate-   S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid sodium salt-   S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid sodium salt monohydrate-   S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid arginine salt 0.25 H₂O-   S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid arginine salt 0.75 H₂O-   8-{4-[2(S)-Amino-propionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid-   8-{4-[2(S)-Amino-propionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid, hydrochloride-   8-{4-[2(R)-Amino-propionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic    acid acetate-   8-{4[(2S)-Methylamino-propionyloxy]piperidin-1-yl}-9-fluoro-(5S)-methyl-6,7    dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid-   8-{4[(2S)-Methylamino-propionyloxy]piperidin-1-yl}-9-fluoro-(5S)-methyl-6,7    dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid,    hydrochloride

Even more preferred compounds of the invention are:

-   S-(−)-9-fluoro-6,7-dihydro-8(-4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid 0.2 hydrate.-   S-(−)-9-fluoro-6,7-dihydro-8(-4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid sodium salt monohydrate.-   8-{4-[2(R)-Amino-propionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]    quinolizine-2-carboxylic acid, acetate.-   8-{4-[2(S)-Amino-propionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]    quinolizine-2-carboxylic acid, hydrochloride.-   S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid arginine salt 0.25 H₂O.-   S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo    [i,j]quinolizine-2-carboxylic acid arginine salt 0.75 H₂O.

Even more preferred compounds of the invention areS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid arginine salt 0.25 H₂O andS-(−)-9-fluoro-6,7-dihydro-8-4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid arginine salt.0.75 H₂O.

In addition to their favourable aqueous solubility, the respectivearginine salts on repeated i.v. administration in rats did not causephlebitis at doses double those of the corresponding sodium salt. Thisfeature would makeS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo [i,j] quinolizine-2-carboxylic acid arginine salt 0.25 H₂O andS-(−)-9-fluoro-6,7-dihydro-8-[4-(hydroxy)-1-piperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid arginine salt 0.75 H₂O suitable forlong term i.v. administration which is commonly undertaken forcritically ill patients or patients in intensive care unit.

The compounds, derivatives, salts, pseudopolymorphs, polymorphs andhydrates of the invention as defined above exhibit the same potentantibacterial activity as S-(−)-Nadifloxacin and also have one or moredesirable physico-chemical properties such as constant moisture content,excellent solubility etc. regardless of the ambient relative humidity,and also have desirable bioavailability and safety profiles.

Generally, conversion of a pharmacologically active compound into a saltor hydrate form induces a change in the compound's physicochemicalproperties such as solubility, absorption velocity, etc. Therefore,study about an effective salt or hydrate form for developing asuccessful new medicine has been conventionally made. Pharmaceuticallymore desirable crystal form may be selected by studying whether or notany polymorphs or pseudopolymorph can be produced and itsphysicochemical properties (see, Remington's Pharmaceutics, Chapter 75Preformulation; Byrn, S. R. Solid Chemistry of Drugs, Academic Press,New York, 1982). The hydrate, one such polymorph or pseudopolymorph, haswater molecules inside the crystal, and thus has a crystalline structuredifferent from that of the anhydrate, as can be verified from theirrespective X-ray diffraction patterns. A polymorph or pseudopolymorphdiffers from the original compound not in its chemical properties, suchas pharmacological activity, but in its physical properties, such ascrystallinity, hygroscopicity, melting point, solubility, solubilizingvelocity, etc. So, the polymorph or pseudopolymorph has been recognisedas pharmaceutically important (see, Morris, KP. et. al. Int. J. Pharm.,108, 15-206 (1994)). In the process of identifying the physico-chemicalproperties of S-(−)-Nadifloxacin, the compound has been found to existas a stable hydrate wherein the proportion of water molecules containedin one molecule varies within a specific range. Here, stability does notmean chemical stability but the difficulty of removing water molecules.That is, a stable hydrate neither loses the water molecule containedtherein, nor absorbs moisture over a wide range of ambient relativehumidity. In contrast, moisture absorption by the anhydrate can varygreatly with the ambient relative humidity. As a result of experimentscarried out by the present inventors,S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid has been shown to exist as a stablehydrate for values of the hydration number z equal to 0.2, 0.5 or 0.75.In addition to the anhydrate S-(−)-Nadifloxacin, the inventors have nowfound thatS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid may exist as a 0.2 hydrate, 0.5 hydrate and 0.75 hydrate crystalforms. Among these, 0.2 is preferred since the change in moisturecontent is the lowest at the hydration number. Although stablehemihydrate and the hemisesquihydrate forms can be prepared, they alteralso to the 0.2 hydrate on vacuum drying at room temperature.

The moisture content of the hydrate varies with the hydration number (z)of the hydrated molecule. The actual moisture content may however,differ from the calculated moisture content depending on differences inrecrystallization conditions, drying conditions, etc. The range of theactual moisture content for the 0.2 hydrate is from 0.9% to 1.1%, eventhough the calculated moisture content is 0.99%.

It has also been found that the relative humidity range at which themoisture content of the anhydrate and the 0.2 hydrate can be maintainedconstant differ from each other. That is, although the anhydrate has aconstant moisture content at a relative humidity of 20% to 75%, the 0.2hydrate is constant at a relative humidity of 20% to 95% (see FIGS. 1and 2).

The hydrates of formula I may be prepared by means of conventionalmethods well known in the art to which the present invention pertains.Particularly, the different hydrates may be prepared merely by changingrecrystallization conditions, and the temperature/vacuum conditionsunder which the crystals are dried. The 0.2 hydrate is prepared bydissolvingS-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid in a minimum volume of organic solvent, preferably acetonitrile orethanol at an elevated temperature, preferably at the reflux temperatureof the solvent and adding an amount of water sufficient to bring aboutcrystallisation after cooling in high yields, filtering and drying theseparated crystals at temperatures up to 40-50° C. for 3-6 hours,preferably 5 hours, in vacuo upto 50 mm of Hg to a constant weight. The0.2 hydrate can also be prepared by dissolvingS-(−)-9-fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid in alkali, preferably 1 molar aqueoussodium hydroxide, heating to 55-60 ° C., acidifying, preferably withconcentrated hydrochloric acid, at 55-60 ° C., maintaining thesuspension at 50-70° C., preferably at 60° C. for at least 30 minutes,cooling, filtering, washing with water and drying the separated crystalsat temperatures up to 40-50 ° C. for 3-6 hours, preferably 5 hours, invacuo upto 50 mm of Hg to a constant weight.

The 0.5 hydrate can be prepared by dissolvingS-(−)-9-fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid in an minimum volume of organic solventsuch as acetone at reflux temperature adding an appropriate amount ofwater at ambient temperature, sufficient to bring about crystallisationafter cooling in high yields, filtering and drying the separatedcrystals at temperatures up to <40° C. for 3-6 hours, preferably 5 hoursto a constant weight.

The 0.75 hydrate can be prepared by suspendingS-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid in water, preferably at 10% (weight byvolume) suspension, formulating into a slurry by vigorous stirringcontinuing stirring at 5° C. for 1-2 hours, adding acetone ca. 5%(weight by volume) with continuation of stirring at 5° C. for 4-5 hours,filtering and drying the product at temperatures <40° C. for 3-6 hours,preferably 5 hours, to a constant weight.

The methods as stated above will be more specifically explained in theexamples described in appropriate later section of this text.

The novel compounds of the Formula I with pharmaceutically acceptablecations are prepared by reacting an appropriate benzoquinolizinecarboxylic acid, for example, S-(−)-nadifloxacin with a base capable ofreleasing the cation X, wherein X is as defined in OR₁ above to give thedesired salt of Formula I. Examples of bases capable of releasing thecation X and examples of reaction conditions are given below.

-   -   a) Salts of the formula I, wherein the cation R1 is lithium,        sodium or potassium are prepared by treating a compound of the        formula I wherein X═OH with LiOH, NaOH, NaHCO₃, Na₂CO₃, KOH,        KHCO₃ or K₂CO₃ in an aqueous or non-aqueous medium.    -   b) Salts of the formula I, wherein the cation R₁ is magnesium,        or calcium, are prepared by treating a compound of the formula I        wherein X═OH with Mg(OH)₂, or Ca(OH)₂, in an aqueous or        non-aqueous medium.    -   c) Salts of the formula I, wherein the cation R₁ is a basic        compound like a basic amino acid or an organic basic amine are        prepared by treating a compound of the formula I, wherein X═OH        with an aqueous or alcoholic solution of the appropriate basic        amino acid or organic basic amine.    -   d) The compound I of the invention which are esters at the        carboxylic acid group may be prepared by treating the free acid        compound I in solution in an appropriate solvent, preferably        N,N-dimethyl formamide, with the corresponding halo compound,        preferably chloro or bromo-compound, in the presence of a base,        preferably anhydrous potassium carbonate, at an elevated        temperature, preferably 50° C. for an extended period of time,        preferably 6 hours.    -   e) The compounds I of the invention which are amides at the        carboxylic acid groups may be prepared by coupling the free acid        compound I with ammonia or an appropriate amine or an amino acid        appropriately protected at the acid functionality of the amino        acids with a protecting group. The —COOH protecting groups for        amino acids are known in the art. Examples of suitable —COOH        protecting groups for amino acids are methyl, ethyl, t-butyl and        benzyl groups. The —COOH protecting group is removed by        hydrolysis or by hydrogenation. The coupling of the —COOH group        of compound I with the amino group of the amino acid is also        known in the art. The reaction may be conducted with or without        a solvent at a range of temperatures in the presence of a        coupling agent.    -   f) The compounds I of the invention which are ethers at the 4-OH        group may be conveniently prepared by condensing the previously        prepared 4-alkoxypiperidine with        S-(−)-diacetoxy-(8,9-difluoro-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxyl)        borane. The reaction may be conducted with or without a solvent        at a range of temperatures in the presence of a condensing        agent.    -   g) The compounds of the invention which are esters at the 4-OH        group may be prepared by treating the free 4-OH compound I with        an organic acid, an organic dibasic acid or appropriate        N-protected amino acid or polypeptide as defined above. Nitrogen        protecting groups are known in the art. Examples of suitable        nitrogen protecting groups are C₁-C₆ acyl, C₂-C₆ alkoxycarbonyl        optionally substituted benzyloxycarbonyl, aryloxycarbonyl,        silyl, trityl, tetrahydropyranyl, vinyloxycarbonyl,        O-nitrophenylsulfonyl, diphenylphosphinyl, p-toluenesulfonyl,        and benzyl. The nitrogen protecting group is removed by methods        known in the art such as hydrogenation or hydrolysis. The ester        forming reaction may be conducted with or without a solvent at a        range of temperatures in the presence of a suitable condensing        agent, known to those skilled in the art.    -   h) The pharmaceutically acceptable acid addition salts of        compounds I are prepared in a conventional manner by treating a        solution or suspension of the free base I with about one        chemical equivalent of a pharmaceutically acceptable acid.        Conventional concentration and recrystalisation techniques are        employed in isolating the salts. Illustrative of suitable acids        are acetic, lactic, oxalic, succinic, maleic, tartaric, citric,        gluconic, ascorbic, benzoic, methanesulfonic, p-toluenesulfonic,        cinnamic, fumaric, phosphoric, hydrochloric, hydrobromic,        hydroiodic, sulfamic, and sulfonic acid.    -   i) The pharmaceutically acceptable cationic salts of compounds I        may be prepared by conventional methods from the corresponding        acids e.g. by reaction with about one equimolar amount of a        base. Examples of suitable cationic salts are those of alkali        metals such as sodium or potassium, alkaline earth metals such        as magnesium or calcium and ammonium or organic amines such as        diethanolamine or N-methylglucamine.

The present invention encompasses a method of treating bacterialinfections, especially resistant Gram-positive organism infections,Gram-negative organism infections, mycobacterial infections andnosocomial pathogen infections in humans and animals, which comprisesadministering systemically or topically to a human or animal in need ofsuch antiinfective therapy an amount of S-(−)-Nadifloxacin or anoptically pure benzoquinolizine carboxylic acid, their derivatives,salts, pseudopolymorphs, polymorphs and hydrates thereof, of the formulaI as defined above, substantially free of its R-(+)-enantiomer, saidamount being sufficient to eradicate such infections. The method avoidsthe concomitant liability of toxic effects associated with theadministration of RS-(±)-isomers by providing an amount ofS-(−)-Nadifloxacin or an optically pure benzoquinolizine carboxylicacid, their derivatives, salts, pseudopolymorphs, polymorphs andhydrates thereof, of the invention, which is insufficient to cause thetoxic effects associated with the racemic mixture of the isomers.

The present invention also encompasses an antiinfective composition forthe treatment of humans and animals in need of therapy for systemic ortopical infections especially resistant Gram-positive organisminfections, Gram-negative organism infections, mycobacterial infectionsand nosocomial pathogen infections, which comprises an amount ofS-(−)-Nadifloxacin and optically pure benzoquinolizine carboxylic acids,the derivatives, salts, pseudopolymorphs, polymorphs and hydratesthereof, of Formula I as defined above, substantially free of theirR-(+)-enantiomers, said amount being sufficient to eradicate saidinfection. The composition should provide a therapeutic dose, which isinsufficient to cause the toxic effects associated with the comparablecompositions comprised of racemic RS-(±)-isomeric mixture.

S-(−)-Nadifloxacin and the compounds of the invention have 2-4 timeshigher antimicrobial activity than RS-(±)-Nadifloxacin againstMupirocin-resistant staphylococci,Methicillin-resistant Staphylococcusaureus (MRSA), Quinolone-resistant Staphylococcus aureus, coagulasenegative staphylococci, such as Methicillin-resistant Staphylococcusepidermidis (MRSE), enterococci, betahemolytic streptococci and viridansgroup of streptococci. The antimicrobial profile of S-(−)-Nadifloxacinand the compounds of the invention have, thus, a potential to addressseveral unmet antibacterial treatment needs ascribed to the mostfrequently encountered Gram-positive bacterial pathogens in clinicalsettings. S-(−)-Nadifloxacin and the compounds of the inventionpossesses superior antibacterial activity against such Gram-positivepathogens which have now become refractory to older first- andsecond-line antibacterials mentioned above (cf. Biological Example 1).Infections such as impetigo, pneumonia, bronchitis, pharyngitis,endocarditis, urinary tract infections and bacteremias caused byStaphylococcus aureus, coagulase negative staphylococci, enterococci,beta haemolytic streptococci and viridans group of streptococci arepotentially amenable to successful treatment with S-(−)-Nadifloxacin andthe compounds of the invention. Intrinsically high potency ofS-(−)-Nadifloxacin and the compounds of the invention coupled with theirpowerful bactericidal action against organisms such as S. aureus,Coagulase negative staphylococci and enterococci rendersS-(−)-Nadifloxacin and the compounds of the invention eminently suitablefor the treatment of infections caused by multi-drug resistant strainsbelonging to this group.

There is a surge of mycobacterial infections due to the spread of AIDSsuch as in several countries of Europe, USA and Asia. AIDS and otherimmunocompromised patients frequently contract mycobacterial infectionsdue to multi-drug resistant M. tuberculosis and other atypicalmycobacteria such as M. intracellulare and M. avium. An embodiment ofthis invention is that the antimycobacterial profile ofS-(−)-Nadifloxacin and the compounds of the invention have been found todisplay significant activity against such organisms and provide avaluable option for the treatment of such problematic diseases. Inaddition, S-(−)-Nadifloxacin and the compounds of the invention havebeen shown by the present inventors to display negligible phototoxicitypotential than the comparator fluoroquinolone drug sparfloxacin. Instudies conducted by the present inventors Sparfloxacin was found to bephototoxic at dosages 25 times lower than S-(−)-Nadifloxacin and thecompounds of the invention.

S-(−)-Nadifloxacin and the compounds of the invention also possesseshigh level of activity against newly emerging Gram-negative pathogenssuch as Chryseobacterium meningosepticum and Chryseobacteriumindologense. These organisms frequently infect immunocompromised adultsas well as premature neonats. These organisms are nosocomial pathogensagainst which most of the currently available antibacterial agentspossess either poor or only borderline activity. The presentinvestigations have shown that S-(−)-Nadifloxacin and the compounds ofthe invention in possessing superior activity against chryseobacteriaand other nosocomial pathogens such as MRSA, enterococci and methicillinsusceptible strains of staphylococci have a potential to becomeexcellent drugs for the treatment of hospital acquired infections (cf.Biological Example 1).

Against bacterial organisms which proliferate in acidic environment suchas the urinary tract, S-(−)-Nadifloxacin and the compounds of theinvention behave in a characteristically different pattern than do theknown fluoroquinolones, such as Ciprofloxcin, Levofloxacin, Ofloxacinand Norfloxacin. In studies carried out by the present inventors theantibacterial potency, that is MIC value, as well as bactericidal actionof S-(−)-Nadifloxacin against Gram-positive pathogens such asstaphylococci and enterococci, and Gram-negative pathogens such as E.coli, Klebsiella, Proteus, Serratia, Citrobacter, and Pseudomonas,unlikethat of Ciprofloxacin and Levofloxacin, is not affected at all by theacidic pH of 5.5. On the contrary, for some organisms the MIC ofS-(−)-Nadifloxacin improves by 100% while that for Ciprofloxacin andLevofloxacin deteriorates in the range of from 50% to 99% (BiologicalExample 2). Further confirmation of these results were obtained bycomparatively evaluating the antibacterial activity ofS-(−)-Nadifloxacin, Ciprofloxacin and Levofloxacin in normal human urineagainst a range of organisms frequently encountered in urinary tractinfections. For Ciprofloxacin and Levofloxacin the loss in antibacterialactivity coupled with the abolition of bactericidal action occurring atacidic pH would lead to recurrent episodes of urinary tract infectionsin patients receiving such fluoroquinolone antibacterial drugs, whereastreatment with S-(−)-Nadifloxacin and the compounds of the invention,would lead to successful and consistent cure, irrespective of the pH orthe nature of the environment in which bacterial pathogens areproliferating. This unpredictable finding with S-(−)-Nadifloxacin is ofgreat clinical relevance and would provide a unique advantage topatients on a regimen of the compounds of the invention for urinarytract infections.

In gram-positive bacteria, especially Staphylococcus strains, resistanceto most of the fluoroquinolones in clinical use is mediated by thepresence of efflux pumps, in particular Nor A efflux pumps, whichaffects the accumulation of the antibiotics within the cell by enhancingefflux, thus preventing the antibiotic action. Current estimates ofprevalence of Nor A bearing strains among ciprofloxacin resistantstaphylococci is about 30-80%.

The present inventors have surprisingly and unexpectedly found that instudies with fluoroquinolone-resistant Staphylococcus strains withefflux pumps, while most of the fluoroquinolones in current clinical usehave shown significantly reduced potency against the efflux-pump bearingStaphylococcus strains, S-(−)-Nadifloxacin, its hydrates, salts,pseudopolymorphs, polymorphs and derivatives thereof, have shown no lossin potency of activity in both in-vitro and in-vivo conditions(Biological Examples 6 & 7). These results support the reducedeffectiveness of the current fluoroquinolones in clinical use intreating infections caused by such efflux-pump-bearing staphylococcalstrains. These results, thus, create a novel opportunity for clinicaluse of the compounds of the invention in treating infections caused byefflux-pump bearing strains, in particular efflux-pump bearingStaphylococcal strains.

The finding of this property of the compounds of the invention is notsuggested by the prior art. In thus behaving differently from thegeneral class of fluoroquinolones, S-(−)-Nadifloxacin and the compoundsof the invention display a property hitherto not yet shown. It hasarisen because of the in-depth studies undertaken by the inventors ofthe compounds of the invention, without any reasonable expectation ofthe kind of result that has been obtained.

The high propensity of S-(−)-Nadifloxacin to display resistance toresistance development in comparison to current fluoroquinolone drugs inclinical use has also now been shown for the first time by theinventors. In studies, which mimic the clinical scenario,S-(−)-Nadifloxacin was evaluated in comparison with trovafloxacin andgatifloxacin by sequential transfer/passages through respective drugcontaining media. Although initially all the three drugs had comparableactivity against S. aureus strain 042, after six passages in drugcontaining media, whilst S-(−)-Nadifloxacin showed a marginal rise of 4%in MIC value, trovafloxacin and gatifloxacin showed approximately 300%and 700% rise respectively in MIC values. This property ofS-(−)-Nadifloxacin to display resistance to resistance development hasconsiderable value clinically. In clinical settings, the ability ofpathogenic bacteria to select a drug resistant variant/subclone whilethe patient is on antibacterial drug therapy often determines theoutcome of the therapy. A drug to which such resistant variant comes upreadily, often witnesses failure of therapy, or a need to increase thedosage significantly, thereby dramatically increasing the chances ofexposing patients to adverse side effects also.

These combined features of S-(−)-Nadifloxacin and optically purebenzoquinolizine carboxylic acids, the derivatives, salts,pseudopolymorphs, polymorphs and hydrates thereof of activity againstbacterial organisms which proliferate in acidic environment, of anability to resist the action of efflux pump present in drug resistantmicrobial strains and of a propensity to display resistance toresistance development, endows the compounds of the invention with aunique clinical potential yet not realised in other fluoroquinoloneantibacterials in current medical use.

The above list of pathogens is merely by way of example and is in no wayto be interpreted as limiting. Examples which may be mentioned ofdiseases, which can be prevented, alleviated and/or cured by theformulations according to the invention are otitis externa, otitismedia; pharyngitis; pneumonia; peritonitis; pyelonephritis; cystitis;endocarditis; systemic infections; bronchitis; arthritis; localinfections; and septic diseases.

These findings have an important implication from the point of view ofthe systemic use of S-(−)-Nadifloxacin and the compounds of theinvention, which in view of their superior potency, superiorbactericidal activity, expanded biospectrum, better bioavailability andimproved tolerability are now enabled to be administered systemically indoses that are insufficient to cause the toxic effects associated withthe administration of racemic RS-(±)-Nadifloxacin and correspondingracemic mixtures of compounds of the invention.

Utilising the substantially optically pure or optically pure isomer ofNadifloxacin or optically pure benzoquinolizine carboxylic acids, thederivatives, salts, hydrates, pseudopolymorphs, or polymorphs thereof,whether in systemic or topical dosage form, results in clearerdose-related definitions of efficacy, diminished toxic effects andaccordingly an improved therapeutic index. It is, therefore, moredesirable to administer the S-(−)-isomer of Nadifloxacin andS-(−)-optically pure benzoquinolizine carboxylic acid, theirderivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof,than RS-(±)-Nadifloxacin and racemic mixtures of compounds of theinvention.

The term “substantially free of its R-(+)-enantiomer” as used hereinmeans that the compositions contain a greater proportion of the S-isomerof Nadifloxacin or an optically pure benzoquinolizine carboxylic acid,their derivatives, salts, pseudopolymorphs, polymorphs and hydratesthereof, in relation to the R-isomer. In a preferred embodiment, theterm “substantially free of its R-isomer” as used herein means that thecomposition is at least 90% by weight of S-(−)-Nadifloxacin or anoptically pure benzoquinolizine carboxylic acid, their derivatives,salts, pseudopolymorphs, polymorphs and hydrates thereof, and 10% byweight or less of the corresponding R-(+)-isomer. In a more preferredembodiment the term “substantially free of the R-enantiomer” means thatthe composition is at least 99% by weight of S-(−)-Nadifloxacin or anoptically pure benzoquinolizine carboxylic acid, their derivatives,salts, pseudopolymorphs, polymorphs and hydrates thereof, and 1% byweight or less of the corresponding R-(+)-isomer. In the most preferredembodiment the term “substantially free of the R-enantiomer” means thatthe composition contains greater than 99% by weight ofS-(−)-Nadifloxacin or an optically pure benzoquinolizine carboxylicacid, their derivatives, salts, pseudopolymorphs, polymorphs andhydrates thereof. These percentages are based on the total amount ofNadifloxacin in the composition. The terms “substantially optically pureS-isomer of Nadifloxacin or an optically pure benzoquinolizinecarboxylic acid, their derivatives, salts, pseudopolymorphs, polymorphsand hydrates thereof” or “substantially optically pure S-Nadifloxacin oran optically pure benzoquinolizine carboxylic acid, their derivatives,salts, pseudopolymorphs, polymorphs and hydrates thereof” and “opticallypure S-enantiomer of Nadifloxacin or an optically pure benzoquinolizinecarboxylic acid, their derivatives, salts, pseudopolymorphs, polymorphsand hydrates thereof” or “optically pure S-Nadifloxacin or an opticallypure benzoquinolizine carboxylic acid, their derivatives, salts,pseudopolymorphs, polymorphs and hydrates thereof” are also encompassedby the above described amounts.

The pharmaceutical compositions are prepared according to conventionalprocedures used by persons skilled in the art to make stable andeffective compositions. In the solid, liquid, parenteral and topicaldosage forms, an effective amount of the active compound or the activeingredient is any amount, which produces the desired results.

For the purpose of this invention the pharmaceutical compositions maycontain the active compounds, S-(−)-Nadifloxacin or an optically purebenzoquinolizine carboxylic acid, their derivatives, salts,pseudopolymorphs, polymorphs and hydrates thereof, in a form to beadministered alone, but generally in a form to be administered inadmixture with a pharmaceutical carrier selected with regard to theintended route of administration and standard pharmaceutical practice.Suitable carriers which can be used are, for example, diluents orexcipients such as fillers, extenders, binders, emollients, wettingagents, disintegrants, surface active agents, humectants, antioxidants,sequestering agents and lubricants which are usually employed to preparesuch drugs depending on the type of dosage form.

Any suitable route of administration may be employed for providing thepatient with an effective dosage of S-(−)-Nadifloxacin or an opticallypure benzoquinolizine carboxylic acid, their derivatives, salts,pseudopolymorphs, polymorphs and hydrates thereof. For example, oral,rectal, parenteral (subcutaneous, intramuscular, intravenous),transdermal, topical and like forms of administration may be employed.Dosage forms include (solutions, suspensions, etc) tablets; pills,powders, troches, dispersions, suspensions, emulsions, solutions,capsules, injectable preparations, patches, ointments, creams, lotions,gels, sprays, shampoos and the like.

Pharmaceutical compositions of the present invention suitable for oraladministration may be presented as discrete units such as capsules,cachets, or tablets, or aerosol sprays, each containing a predeterminedamount of the active ingredient, as a powder or granules, or as asolution or a suspension in an aqueous liquid, a non-aqueous liquid, anoil-in-water emulsion, or a water-in-oil liquid emulsion. Suchcompositions may be prepared by any of the methods of pharmacy, but allmethods include the step of bringing into association the activeingredient with the carrier which constitutes one or more necessaryingredients. In general, the compositions are prepared by uniformly andintimately admixing the active ingredient with liquid carriers or finelydivided solid carriers or both, and then, if necessary, shaping theproduct into the desired presentation.

The compositions of the present invention include compositions such assuspensions, solutions, elixirs, aerosols, and solid dosage forms.Carriers as described in general above are commonly used in the case oforal solid preparations (such as powders, capsules and tablets), withthe oral solid preparations being preferred over the oral liquidpreparations. The most preferred oral solid preparation is tablets.

Because of their ease of administration, tablets and capsules representthe most advantageous oral dosage unit form, in which case solidpharmaceutical carriers are employed. Examples of suitable carriersinclude excipients such as lactose, white sugar, sodium chloride,glucose solution, urea, starch, calcium carbonate, kaolin, crystallinecellulose and silicic acid, binders such as water, ethanol, prepanol,simple syrup, glucose, starch solution, gelatin solution, carboxymethylcellulose, shellac, methyl cellulose, potassium phosphate and polyvinylpyrrolidone, disintegrants such as dried starch, sodium alginate, agarpowder, laminaria powder, sodium hydrogen carbonate, calcium carbonate,Tween (fatty acid ester of polyoxyethylenesorbitan), sodium laurylsulfate, stearic acid monoglyceride, starch, and lactose, disintegrationinhibitors such as white sugar, stearic acid glyceryl ester, cacaobutter and hydrogenated oils, absorption promoters such as quaternaryammonium bases and sodium lauryl sulfate, humectants such as glyceroland starch, absorbents such as starch, lactose, kaolin, bentonite andcolloidal silicic acid, and lubricants such as purified talc, stearicacid salts, boric acid powder, polyethylene glycol and solidpolyethylene glycol.

The tablet, if desired, can be coated, and made into sugar-coatedtablets, gelatin-coated tablets, enteric-coated tablets, film-coatedtablets, or tablets comprising two or more layers.

If desired, tablets may be coated by standard aqueous or nonaqueoustechniques.

Desirably, each tablet, cachet, capsule contains from about 200 mg toabout 1500 mg of the active ingredient. Most preferably, the tablet,cachet or capsule contains either one of three dosages, about 200 mg,about 400 mg, or about 600 mg of the active ingredient.

In molding the pharmaceutical composition into pills, a wide variety ofconventional carriers known in the art can be used. Examples of suitablecarriers are excipients such as glucose, lactose, starch, cacao butter,hardened vegetable oils, kaolin and talc, binders such as gum arabicpowder, tragacanth powder, gelatin, and ethanol, and disintegrants suchas laminaria and agar.

In molding the pharmaceutical composition into a suppository form, awide variety of carriers known in the art can be used. Examples ofsuitable carriers include polyethylene glycol, cacao butter, higheralcohols, gelatin, and semi-synthetic glycerides.

A second preferred method is parenterally for intramuscular, intravenousor subcutaneous administration.

A third preferred route of administration is topically, for whichcreams, ointments, sprays, shampoos, lotions, gels, dusting powders andthe like are well suited. Generally, an effective amount of the compoundaccording to this invention in a topical form is 0.1% to about 10% byweight of the total composition. Preferably, the effective amount is 1%by weight of the total composition.

For topical application, there are employed as non-sprayable forms,viscous to semi-solid or solid forms comprising a carrier compatiblewith topical application and having a dynamic viscosity preferablygreater than water. Suitable formulations include but are not limited tosolutions, suspensions, emulsions, creams, ointments, powders,liniments, salves, aerosols, etc., which are, if desired, sterilized ormixed with auxiliary agents, e.g. preservatives, antioxidants,stabilizers, wetting agents, buffers or salts for influencing osmoticpressure, etc. For topical application, also suitable are sprayableaerosol preparations wherein the active ingredient preferably incombination with a solid or liquid inert carrier material.

In addition to the common dosage forms set out above, the compounds ofthe present invention may also be administered by controlled releasemeans and/or delivery devices such as those described in U.S. Pat. Nos.3,845,770; 3,916,899; 3,536,809; 3,598,123 and 4,008,719; thedisclosures of which are hereby incorporated by reference.

When the pharmaceutical composition is formulated into an injectablepreparation, in formulating the pharmaceutical composition into the formof a solution or suspension, all diluents customarily used in the artcan be used. Examples of suitable diluents are water, ethyl alcohol,polypropylene glycol, ethoxylated isostearyl alcohol, polyoxyethylenesorbitol, and sorbitan esters. Sodium chloride, glucose or glycerol maybe incorporated into a therapeutic agent.

The antimicrobial pharmaceutical composition may further containordinary dissolving aids, buffers, pain-alleviating agents, andpreservatives, and optionally coloring agents, perfumes, flavors,sweeteners, and other drugs.

A specific embodiment of the invention is the preparation of storagestable compositions of the compounds of the invention of formula I. Suchstable compositions can be advantageously made through the use ofselective stabilizers. Different stabilizers are known to those skilledin the art of making pharmaceutical compositions. Of special utility formaking storage stable compositions of the compound of the invention offormula I, stabilizers such as disodium EDTA, tromethamine,cyclodextrins such as gamma-cyclodextrin, beta-cyclodetrin,hydroxy-propyl-gamma-cyclodextrin have been found to be useful.

A specific embodiment of the invention utilises arginine as an excipientin compositions to facilitate the aqueous solubility of the compounds ofthe invention which comprises utilising an appropriate molar amount ofarginine with a specific compound of the invention. For example, a 0.7molar amount of arginine added to a molar amount ofS-(−)-9-fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5Hbenzo[i,j] quinolizine-2-carboxylic acid arginine salt 0.75 H₂O, raisesthe aqueous solubility of the salt from 94 mg/ml to a value >200 mg/ml.

In a specific embodiment of the invention, the pharmaceuticalcompositions contain an effective amount of the active compounds ofS-(−)-9-fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5Hbenzo[i,j]quinolizine-2-carboxylic acid (also called S-(−)-Nadifloxacin)or one of the specific optically pure derivatives, salts,pseudopolymorphs, polymorphs or hydrates thereof described in thisspecification in admixture with a pharmaceutically acceptable carrier,diluent or excipients, and optionally other therapeutic ingredients.

The prophylactic or therapeutic dose of S-(−)-Nadifloxacin and opticallypure benzoquinolizine carboxylic acids, the derivatives, salts,pseudopolymorphs, polymorphs or hydrates thereof, in the acute orchronic management of disease will be calculated based on theprophylactic or therapeutic dose ofS-(−)-9-fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7dihydro-1-oxo-1H,5Hbenzo[i,j] quinolizine-2-carboxylic acid and will vary with the severityof condition to be treated, and the route of administration. Inaddition, the dose, and perhaps the dose frequency, will also varyaccording to the age, body weight and response of the individualpatient. In general, the total daily dose range, for S-(−)-Nadifloxacinor an optically pure benzoquinolizine carboxylic acids, the derivatives,salts, pseudopolymorphs, polymorphs or hydrates thereof, for theconditions described herein, is from about 200 mg to about 1500 mg, insingle or divided doses. Preferably, a daily dose range should bebetween about 400 mg to 1200 mg, in single or divided dosage, while mostpreferably a daily dose range should be between about 500 mg to about1000 mg in divided dosage. While intramuscular administration may be asingle dose or upto 3 divided doses, intravenous administration caninclude a continuous drip. It may be necessary to use dosages outsidethese ranges in some cases as will be apparent to those skilled in theart. Further, it is noted that the clinician or treating physician willknow how and when to interrupt, adjust, or terminate therapy inconjunction with individual patient's response. The term “an amountsufficient to eradicate such infections but insufficient to cause saidtoxic effect” is encompassed by the above-described dosage amount anddose frequency schedule.

The invention is further defined by reference to the following examplesdescribing in detail the preparation of the composition of the presentinvention as well as their utility. It will be apparent to those skilledin the art that many modifications, both to materials and methods may bepracticed without departing from the purpose and scope of thisinvention.

The following examples illustrate the methods of preparation of thecompounds of the invention and are provided only as examples, but not tolimit the scope of the compounds of the invention.

EXAMPLE 1S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolzine-2-carboxylic acid anhydrate

Method A

S-(−)-9-fluoro-6,7dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid (3.0 g) obtained according to theprocess described in literature [K. Hashimoto et. al., Chem. Pharm.Bull. 44, 642-5(1996)] was dissolved in acetonitrile (250 ml) at 85° C.The resulting clear solution was filtered (to remove if any fibrousmaterial is in suspension). The filtrate was concentrated to 125 ml andleft at room temperature for crystallization. The crystals thusseparated were filtered and dried in a drying cabinet at 40° C. for 2 hrin vacuum at 50 mm of Hg to obtain constant weight. Yield 2.6 g (86%).

Method B:

S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid (2.0 g) obtained according to theprocess described in literature [K. Hashimoto et.al., Chem. Pharm. Bull.44, 642-5(1996)] was dissolved in ethyl alcohol (95%; 200 ml) at 80° C.The obtained clear solution was filtered (to remove if any fibrousmaterial is in suspension), concentrated to 100 ml and left forcrystallization. The separated solid was filtered and dried in a dryingcabinet at 40° C. for 3 hr in vacuum at 50 mm of Hg to obtain constantweight. Yield 1.7 g (85%).

M.p. 258-62° C., moisture content 0% (by Karl Fisher method) [α]_(D)²⁶−299°, HPLC purity 99.8%

The X-ray diffraction pattern and the DSC analysis of the sample wereidentical to that of the anhydrate shown in FIG. 3 and FIG. 7respectively.

EXAMPLE 2S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid 0.2 hydrate

Method A:

S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid (5.0 g) was dissolved inacetonitrile (500 ml) at 100° C. and filtered to remove suspendedfibrous impurities. Distilled water (1500 ml) was added. On standingovernight at 5° C., the solid separated was filtered and dried at <50°C. for 5 hrs in vacuum at 50 mm of Hg to obtain constant weight. Yield3.5 g (70%).

Method B:

S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid (4.0 g) was dissolved in ethylalcohol (200 ml) at 85° C. to obtain clear solution and distilled water(700 ml) was added. On standing overnight at 5° C., the solid thusseparated was filtered and dried at <50° C. for 5 hrs in vacuum at 50 mmof Hg to obtain constant weight. Yield 3.1 g (77%).

Method C:

S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid (1.0 g) was dissolved in aqueousNaOH (1M, 10 ml) with stirring at room temperature, and filtered toremove suspended fibrous impurities. The obtained clear solution washeated at 55-60° C. with stirring for 15 min and acidified with 35% HCl(1.5 ml) in hot. The suspension was stirred at 50-70° C. preferably at60° C. for at least 30 min, cooled at room temperature, filtered andwashed with water (10 ml) to furnish the hydrate. The obtained hydratewas dried at <50° C. for 3 hrs in vacuum at 50 mm of Hg to obtainconstant weight. Yield 0.7 g (70%).

M.p. 248-52° C., moisture content 0.9-1.04% (by Karl Fisher method),[α]_(D) ²⁶−259.75° and HPLC 99.74%.

The X-ray diffraction pattern and the DSC analysis of the sample wereidentical to that of the 0.2 hydrate shown in FIG. 4 and FIG. 8respectively.

EXAMPLE 3S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid 0.5 hydrate (hemihydrate)

S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid (8.0 g) was suspended in acetone (400 ml)and refluxed to obtain a clear solution. Heating was stopped and water(1500 ml) was added. The obtained mixture was kept over night at 5° C.The solid thus separated was filtered, washed with chilled acetone (5ml) and dried at room temperature for 48 hr to obtain constant weight.Yield 6.2 g (77.5%).

M.p. 256-58° C., moisture content 2.42% (by Karl Fisher method), HPLCpurity 99.34% and [α]_(D) ²⁶−260°.

The X-ray diffraction pattern and the DSC analysis of the sample wereidentical to that of the hemihydrate shown in FIG. 5 and FIG. 9respectively.

EXAMPLE 4S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid 0.75 hydrate (hemisesquihydrate)

S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-[-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid (10.0 g) was suspended in water (100 ml)and formulated into a slurry over a period of at least 1 hr withvigorous stirring. The obtained slurry was stirred at 5° C. for 1hr,acetone (200 ml) was added and stirring continued at 5° C. for at least4 hr. The solid thus separated was filtered, washed with chilled acetone(5 ml) and dried at room temperature for 24 hr to obtain constantweight. Yield 2.95 g (30%).

M.p. 256-58° C., moisture content 3.294% (by Karl Fisher method) HPLCpurity 99.44% and [α]_(D) ²⁶−253°.

The X-ray diffraction pattern and the DSC analysis of the sample wereidentical to that of the hemisesquihydrate shown in FIG. 6 and FIG. 10respectively.

EXAMPLE 5S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid, sodium salt monohydrate

Method A:

S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid (1.0 -g, 2.777 mmole) was dissolvedin acetonitrile (100 ml) at 90° C. to obtain clear solution, aqueousNaOH (2.67 ml, 1.04 mole) was added dropwise, the mixture was refluxedfor 30 min and allowed to cool at room temperature for crystallization.The crystals thus separated were filtered and dried at 50° C. for 3 hrin vacuum at 50 mm of Hg to obtain constant weight. Yield 0.86 g (81%).

Method B:

The experiment was repeated using acetone in place of acetonitrile togive the same product.

Method C:

Aqueous sodium hydroxide solution (1N, 1.39 ml, 1.39 mmol) was added tothe stirred powder ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid (0.5 g, 1.39 mmol) and diluted with water (10 ml). The resultingsolution was stirred for 30 min., passed through micro filter and freezedried to furnishS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid sodium salt monohydrate. Yield 0.54 g (98%).

M.p. 285° C. (dec), m/z 383 (M+H), [α]_(D) ²¹−261° (1% water solution),solubility >1000 mg/ml in water, PMR (D₂O) δppm: 1.29 (3H, d, j=7.0 Hz,CH₃), 1.43-1.65 (2H, m, H₆), 1.65-2.1 (4H, m, H₃′, and H_(5′)), 2.6-3.2(6H, m, H₂′, H_(6′) and H₇), 3.71 (1H, m, H_(4′)), 4.47 (1H, m, H₅),7.63 (1H, d, J=16.5 Hz, H₁₀), 8.32 (1H, S, H₃), moisture content 5.06%(by Karl Fisher method) and HPLC purity 98.7%.

EXAMPLE 6S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid potassium salt monohydrate

Aqueous potassium hydroxide solution (0.5%, 15.6 ml, 1.39 mmol) wasadded to the stirred powder ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid (0.5 g, 1.39 mmol). The resultingsolution was stirred for 30 min., passed through micro filter and freezedried to provideS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid potassium salt. Yield 0.568 g (99%), m.p. >300° C., m/z 399 (M+H),[α]_(D) ²⁵−255° (1% water solution), solubility >1000 mg/ml in water,PMR (DMSO-d₆) δppm: 1.3 (3H, d, j=6.8 Hz, CH₃), 1.4-1.7 (2H, m, H₆),1.7-1.95 (2H, m, H_(3′) and H_(5′)), 1.95-2.2 (2H, m, H_(3′) andH_(5′)), 2.75-2.98(2H, m, H_(2′) and H₇), 2.98-3.3 (4H, m, H₂′, H₆′, andH₇), 4.1 (1H, m, H_(4′)), 4.5 (1H, m, H₅), 7.84 (1H, d, J=12.6 Hz, H₁₀)8.3 (1H, S, H₃).

EXAMPLE 7S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid, L-arginine salt 0.25 hydrate

Aqueous L-arginine solution (1%, 24.2 ml, 1.39 mmol) was added to thestirred solution ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid (0.5 g, 1.39 mmol) in methanol (20ml). The resulting solution was stirred for 30 min., passed through amicro filter and concentrated to dryness to furnishS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid, L-arginine salt. Yield 0.7 g(93.4%), m.p. 255-60° C., m/z 535 (M+H), [α]_(D) ²⁵−193.3° (1% methanolsolution) solubility >75 mg/ml in water, PMR (D₂O) δppm: 1.32 (3H, d,j=6.8 Hz, CH₃), 1.5-1.7 (2H, m, H₆), 1.7-2.2 (8H, m, H_(3′) H_(5′) and2XCH₂), 2.7-3.3 (8H, m, H_(2′), H_(6′), H₇ and NCH₂), 3.5(1H, m, CH),3.75(1H, m, H_(4′)), 4.5 (1H, m, H₅), 7.85 (1H, d, J=12.6 Hz, H₁₀) 8.5(1H, S, H₃).

EXAMPLE 8S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid, L-arginine salt 0.75 hydrate

L-(+)-Arginine (0.958 g., 5.5 mmoles) was added in portions to asuspension solution ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid 0.2 hydrate (2.0 g., 5.5 mmole) inmethanol (400 ml). The obtained solution was concentrated in vacuum togive the desired product as a yellow solid, which was dried at 50° C. at50 mm/Hg for 5 hours. Yield 3.0 g. (100%), m.p. 220-223 ° C. (dec), m/z535 (M+H), moisture content 2.3% (by Karl Fisher, required 2.46%),[α]_(D) ²⁵−144° (1% methanol c=1), solubility 93 mg/ml.

EXAMPLES 9 & 10

Similarly prepared wereS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid L-lysine salt monohydrate, andS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid L-histidine salt 0.2 hydrate.

S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid L-lysine salt monohydrate

Yield 0.7 g (99%), m.p. 235-40° C., m/z 506 (M+H), [α]_(D) ²⁵−177° (1%methanol solution) solubility 75 mg/ml in water. PMR (DMSO-d₆) δppm:1.38 (3H, d, j=6.8 Hz, CH₃), 1.48-2.25 (10H, m, H_(3′) H₅′, H₆ and2×CH₂), 2.5-2.83 (4H, m, 2×CH₂), 2.85-3.4 (6H, m, H_(2′), H_(6′), andH₇), 3.72-3.88 (2H, m, H_(4′) and CH), 4.75 (1H, m, H₅), 7.78 (1H, d,J=12.6 Hz, H₁₀), 8.8 (1H, S, H₃).

S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid L-histidine salt 0.2 hydrate

Yield 0.67 g (94%), m.p. 270-80° C., m/z 515 (M+H), [α]_(D) ²⁵−216° (1%methanol solution) solubility 75 mg/ml in water, PMR (D)MSO-d₆) δppm:1.42 (3H, d, j=6.8 Hz, CH₃), 1.48-1.70 (2H, m, H₆), 1.75-2.23 (4H, m,H_(3′) and H_(5′)), 2.78-3.31 (6H, m, H_(2′), H_(6′), and H₇), 3.5 (1H,m, CH), 3.71 (2H, m, CH₂), 4.15 (1H, m, H_(4′)), 4.78 (1H, m, H₅), 6.9(1H, s, imidazole H), 7.62 (1H, s, imidazole H), 7.83 (1H, d, J=12.5 Hz,H₁₀) 8.86 (1H, S, H₃).

EXAMPLE 11 PivaloyloxymethylS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate

S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid (1.0 g, 2.77 mmol) was dissolved inN,N-dimethyl formamide (25 ml) and solution was stirred at 50° C.Powdered anhydrous potassium carbonate (0.385 g, 2.77 mmol) was added tostirred solution and stirring was continued for 6 hr at 50° C. Chloromethyl pivalate (2.0 g, 13.88 mmol) was added to the resulting mixtureand stirred for 40 h at 50° C. The reaction mixture was concentrated,triturated with water, extracted with chloroform to give crude product,which was purified by chromatography. Yield 0.9 g (71%), m.p 198-200°C., m/z 475(M+H).

EXAMPLE 12

Similarly prepared to the product of Example 11 were AcetoxymethylS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate.Chloromethyl acetate was used in place of chloromethyl pivalate. Yield0.35 g (56%), m.p. 180° C., m/z 433(M+1), [α]_(D) ^(22.5)−251° (1% CHCl₃solution).

EXAMPLE 13

Similarly prepared to the product of Example 11 was PivaloyloxyethylS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate. Chloroethyl pivalate was used in placeof chloromethyl pivalate. Yield 0.08 g (59%), m.p. 92-95° C., m/z489(M+1), [α]_(D) ^(22.5)−174.5° (0.4% methanol solution).

EXAMPLE 14

Similarly prepared to the product of Example 11 was PropionoxymethylS-(−)-9-fluoro-6,7-dihydro-8-(4-hyroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylate. Bromoethyl acetate was used in place ofchloromethyl pivalate. Yield 0.4 g (67%), m.p. 185-187° C., m/z 447(M+1), [α]_(D) ^(22.5)−186° (1% chloroform solution).

EXAMPLE 15 CarboxymethylS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylate(sodium salt)

S-(−)9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid (0.72 g, 2 mmol) was dissolved inN,N-dimethyl formamide (25 ml) and solution was stirred at 50° C.Powdered anhydrous potassium carbonate (0.385 g, 2.77 mmol) was added tostirred solution and stirring was continued for 6 hr at 50° C. Bromoacetic acid t-Butyl ester (1.9 g, 10 mmol) was added to the resultingmixture and stirred for 40 h at 50° C. The reaction mixture wasconcentrated, triturated with water, extracted with chloroform to givecrude product, which was purified by chromatography. Yield 0.76 g (80%).

The t-Butyl group was removed by treatment with trifluoroacetic acid toget the desired product.

EXAMPLE 16S-(−)-9-fluoro-6,7-dihydro-8-(4-methoxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid

A mixture ofS-(−)-diacetoxy-(8,9-difluoro-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxyl)borane(0.2 g, 0.49 mmol) and 4-methoxypiperidine (0.226 g, 1.9 mmol) inacetonitrile (8 ml) was stirred at 100° C. for 24 h. The reactionmixture was concentrated, triturated with water and filtered. Theobtained solid was dissolved in acetonitrile (8 ml), treated with 1Naqueous NaOH solution (10 ml) and stirred to obtain a clear solution.The resulting solution was acidified with conc. HCl, the separatedprecipitate was filtered, washed with water and dried. The obtainedcrude product was purified by chromatography. Yield 0.07 g (38%), m.p194° C., m/z 375(M+H), [α]_(D) ²⁶−209.75° (0.5% methanol solution), PMR(CDCl₃) δppm: 1.55 (3H, d, j=6.8 Hz, CH₃), 1.8-1.9 (2H, m, H₆), 1.9-2.3(4H, m, H_(3′) and H_(5′)), 2.8-3.2 (6H, m, H_(2′), H_(6′) and H₇), 3.25(1H, m, H_(4′)), 3.45 (3H, s, CH₃), 4.55 (1H, m, H₅), 8.2 (1H, d, J=16.5Hz, H₁₀), 8.7 (1H, S, H₃), 15.1 (1H, bs, COOH, D₂O exchangeable).

EXAMPLE 17S-(−)-9-fluoro-6,7-dihydro-8-(4-acetoxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine2-carboxylic acid

Acetic anhydride (0.312 g, 3.6 mmol) was added to a stirred mixture ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid (0.65 g, 1.8 mmol) and N,N4-dimethylaminopyrdine (0.01 g) inpyridine (10 ml), stirring was continued for 3 h at ambient temperature.The reaction mixture was concentrated, triturated with water, filtered,washed with water and dried. The obtained crude product was purified bychromatography. Yield 0.69 g (95%), m.p 230-35° C., m/z 403(M+H),[α]_(D) ²⁵−239°0 (1% methanol solution), PMR (CDCl₃) δppm: 1.51 (3H, d,j=6.8 Hz, CH₃), 1.85-1.9 (2H, m, H₆), 2.1 (3H, s, COCH₃), 1.9-2.3 (4H,m, H_(3′) and H_(5′)), 2.9-3.4 (6H, m, H_(2′), H_(6′) and H₇), 4.5 (1H,m, H₅), 5.0 (1H, m, H_(4′)), 8.2 (1H, d, J=16.5 Hz, H₁₀), 8.7 (1H, S,H₃) 15.1 (1H, bs, COOH, D₂O exchangeable).

EXAMPLE 18S-(−)9fluoro6,7-dihydro-8-(4-pivaloyloxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolzine-2-carboxylic acid.

Pivaloyl chloride (0.08 g, 0.66 mmol) was added to a stirred mixture ofS-(−)-9-fluoro-6,7-dihydro-8-4hydroxypiperidin-1-yl)5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid (0.08 g, 0.22 mmol) and N,N4dimethylaminopyridine (0.005 g) inpyridine (5 ml), stirring was continued for 3 h at ambient temperature.The reaction mixture was concentrated, triturated with water, filtered,washed with water and dried. The obtained crude product was purified bychromatography. Yield 0.05 g (50%), m.p 200-05° C., m/z 445(M+H),[α]_(D) ²⁶−199.5° (0.5% methanol solution), PMR (CDCl₃) δppm: 1.26 (9H,S, 3× CH₃), 1.57 (3H, d, j=6.8 Hz, CH₃), 1.8-1.95 (2H, m, H₆), 1.95-2.36(4H, m, H_(3′) and H_(5′)), 2.9-3.5 (6H, m, H_(2′), H_(6′) and H₇), 4.58(1H, m, H₅), 5.05 (1H, m, H_(4′)) 8.22 (1H, d, J=16.5 Hz, H₁₀), 8.74(1H, S, H₃).

EXAMPLE 19S-(−)-9-fluoro-6.7-dihydro-8-(4-[(β-D-tetraacetylglucopyranosyl)oxy]-piperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid

A solution of acetobromoglucose (1.71 g., 4.16 mmol) in dichloroethane(20 ml.) was added toS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid (1.0 g, 2.77 mmol), 4° A molecularsieves (˜100 g) and silver carbonate (3.066 g., 11.11 mmol). Thereaction mixture was heated at 60-70° C. under argon atmosphere for 20hrs. in the dark. The solids were filtered off and the filtrate wasconcentrated in vacuum to give the product. Column chromatography of thecrude product gave the tetraacetate. Yield 1.75 g. m.p 157-158° C., m/z691(M+H), [α]_(D) ²⁶−199.5° (0.5% methanol solution), PMR (CDCl₃) δppm:1.42 (3H, d, j=6.8 Hz, CH₃), 1.73-1.98 (2H, m, H₆), 2.01 (3H, s, COCH3)2.20 (9H, s, 3xCOCH3), 2.60-3.40 (8H, m), 3.80-4.52 (7H, m), 5.46-5.42(4H, m) 5.98 (1H, d, J=10.1 Hz), 7.91(1H, d, J=12.5 Hz), 8.77 (1H, s).

EXAMPLE 20S-(−)9-fluoro6,7-dihydro-8-(4[(β-D-glucopyranosyl)oxy]-piperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid

S-(−)-9fluoro-6,7-dihydro-8(4-[(P-D-tetraacetylglucopyranosyl)oxy]-piperidin-1-yl)5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid (0.144 g., 0.20 mmol) was dissolved in a 3:1 mixture of methanoland water. Lithium hydroxide (87 mg., 2.00 mmol) was added to thesolution and the mixture was stirred at room temperature for 0.5 hr. Themixture was evaporated to dryness and the residue was dissolved in smallamount of methanol, filtered and the filtrate was evaporated to give theproduct.

EXAMPLES 21 TO 23 General method for making amides ofS-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid with an amino acid e.g. glutamic acid9-fluoro-8(4-hydroxypiperidin-1-yl)(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-[2(S)-amino-1,5-pentanedioicacid] carboxamide, disodium salt

S-(−)-9-Fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid (360 mg, 1.0 mmol) and triethylamine(0.145 ml, 1.0 mmol) were dissolved in dimethylacetamide (15 ml).Isobutylchloroformate (0.13 ml, 1.0 mmol) was added under ice coolingand stirred for 5 min. A solution of S-glutamic acid dimethyl esterhydrochloride (422 mg, 2.0 mmol) and triethylamine (2 mmol, 0.28 ml) indimethylacetamide (10 ml) was added, followed by addition of4-(dimethylamino) pyridine (125 mg, 1.0 mmol) and the mixture wasstirred at RT overnight. The reaction mixture was diluted with ethylacetate (50 ml), washed with 0.5 N HCl, saturated NaHCO₃ solution,brine, dried (Na₂SO₄) and evaporated under vacuum. The residue wasdissolved in methanol (10 ml), added 1N NaOH (1.1 ml) and stirred for2-3 h at RT. The reaction mixture was concentrated, acidified with 1NHCl and dissolved in ethyl acetate (50 ml). The organic layer was washedwith brine, dried (Na₂SO₄), solvent was evaporated, residue purified bycolumn chromatography and freeze dried to give the free acid. Yield (150mg, 30%). Dissolved the free acid (150 mg, 0.3 mmol) and NaHCO₃ (50 mg,0.6 mmol) in water and freeze dried to give the product.

Similarly made were amides using the amino acids such as alanine andhistidine.

EXAMPLES 24-37

General method for making the amino acid esters of the 4′ hydroxy ofpiperidinyl moiety ofS-(−)-9-Fluoro-8(4-hydroxypiperidin-1-yl)-(5S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid, e.g. the lysine ester.

8-{4-2(S),6-Diaminohexanoyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid, dihydrochloride

S(−)-9-Fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid (360 mg, 1.0 mmol) and triethylamine (0.14ml, 1.0 mmol) were dissolved in dimethylacetamide (15 ml).Bis-t-butyloxycarbonyl-S-lysine (415 mg, 1.2 mmol) and4-(dimethylamino)pyridine (150 mg, 1.2 mmol) were added, followed by theaddition 1,3-dicyclohexylcarbodiimide (206 mg, 1.0 mmol) under icecooling. The reaction mixture was stirred for 30 minutes at 0° C.followed by overnight stirring at RT. The reaction mixture was filtered,diluted with ethyl acetate, washed with 0.5 N HCl, saturated NaHCO₃solution, brine, dried (Na₂SO₄) and evaporated to give residue. Theresidue was treated with trifluoroacetic acid (10 ml), stirred themixture at RT for 30 min. and evaporated under reduced pressure. Theresidue was triturated with ether to give the precipitated. Theprecipitates were further purified by column chromatography, dissolvedin 0.1 N HCl and freeze dried to give the product. Yield (374 mg, 69%)

Similarly esters were made with the following amino acids: S-Ala,S-Ala-S-Ala, R-Ala, R-Ala-R-Ala, N-Methyl S-Ala, S-Leu, R-Leu, S-Phe,S-Pro, S-Asp, Nitro-S-Arg, S-Arg, Nitro-S-Arg-Nitro-S-Arg, S-Arg-S-Arg,The data corresponding to the respective compounds made are provided inthe following table:

Amino acid linked Melting at 4-OH group of Point MASS HPLC MoistureExample 1 compound Salt Yield % ° C. (M + H) Purity (%) Content 25.S-Ala HCl, H₂O 95   160-5(d) 432 99.0 3.4 26. R-Ala HCl, H₂O 92 225 43299.0 4.7 27. R-Ala-S-Ala HCl, 0.5 H₂O 90 190-93 503 97.0 1.1 28. R-LeuHCl, H₂O 94 220-33 474 99.0 3.4 29. N-Me-S-Ala HCl, 0.5 H₂O 91 140-50446 97.5 1.9 30. R-Ala AcOH 98 125-27 432 99.7 — 31. S-Val HCl, 0.75 H₂O93 160-61 460 96.2 2.7 32. S-Ala-S-Ala HCl 60 175-80 503 97.8 — 33.R-Ala-R-Ala HCl 75  95-100 503 98.0 — 34. S-Arg (Nitro) HCl 70 113-16588 93.0 — 35. S-Arg HCl 70 178-82 603 94.0 — 36. [S-Arg(Nitro)]₂ HCl37. S-Arg-S-Arg 3HCl

BIOLOGICAL EXAMPLES

Microbiological and pharmacological studies can be used to determine therelative potency, and the profile of specificity of the optically pureenantiomers, and the racemic mixture of Nadifloxacin as antibacterialagent with a spectrum of activity as described in the specificationabove.

BIOLOGICAL EXAMPLE 1

In-vitro Antimicrobial Activity Test

The activity of the compounds of the invention in vitro can beillustrated as follows:

The comparative antimicrobial activity of S-(−)-Nadifloxacin,RS-(±)-Nadifloxacin, Mupirocin and Levofloxacin against variousmicroorganisms is given in Table 1. The test method was in accordancewith the standard NCCLS protocol.

TABLE 1 Comparative MICs (μg/ml) of S-(−)-Nadifloxacin,RS-(±)-Nadifloxacin, Mupirocin and Levofloxacin S-(−)- RS-(±)- LEVO-ORGANISMS NADIFLOXACIN NADIFLOXACIN MUPIROCIN FLOXACIN Staphylococcusaureus ATCC 25923 0.025 0.05 0.4 0.2 MRSA STA-4 0.4 0.8 0.4 >12.5 MRSESTE-22 0.4 1.56 0.2 >12.5 Mupirocin-resistant Staph STA-34 0.4 1.56 >40012.5 Propioni bacterium acnes 0.1 0.2 >1000 1.0 Streptococcus pneumoniaeATCC 6303 0.2 0.4 0.2 0.8 Streptococcus pyogenes 0.2 0.4 0.025 0.4Viridans group Streptococci 0.2 0.4 0.2 1.56 Enterococcus faecalis ATCC29212 0.2 0.4 >12.5 0.8 Enterococcus faecium 0.4 0.8 >0.8 1.56Corynebacterium jeikeium 0.05 0.2 >12.5 0.4 Haemophilus influenzae 0.0250.05 N.A. 0.03 Escherichia coil ATCC 25922 0.2 0.8 N.A. 0.05 Serratiamarcescens 0.4 1.56 N.A. 0.1 Pseudomonas aeruginosa 1.56 3.12 N.A. 3.12Bacteroides fragilis 0.8 3.12 N.A. 6.25 Mycobacterium tuberculosis ATCC0.8 1.56 — 0.4 27294 Mycobacterium intracellulare 1.56 3.12 — 0.8Mycobacterium avium 3.12 6.25 — 12.5 Chryseobacterium meningosepticum0.8 1.56 — 6.25 1. Mupirocin resistant MRSA strains with very high MICsof >400 μg/ml can effectively be inhibited by S-(−)-Nadifloxacin orracemic Nadifloxacin at much lower MICs of 0.4-1.56 μg/ml. For suchstrains, levofloxacin is 30 times less active than S-(−)-Nadifloxacinand 8 times less active than RS(±)Nadifloxacin. 2. S-(−)-Nadifloxacinhas 2-4 times higher activity than racemic-Nadifloxacin. 3.S-(−)-Nadifloxacin has 48 times higher activity than levofloxacinagainst nosocomial pathogens like enterococci and chryseobacteriummeningosepticum.

BIOLOGICAL EXAMPLE 2

Effect of pH on Potency of Fluoroqinolones

The test method was in accordance with the standard NCCLS protocolemploying test media adjusted at pH 5.5 and 7.0.

TABLE 2 Effect of pH at 5.5 on % loss (−)/gain (+) in potency ofFluoroquinolones against Urinary Tract Pathogens % CHANGE IN POTENCY ATpH 5.5* ORGANISM S-(−)-Nadi Cipro Levo S. aureus 25923 +100 −75 −75 S.aureus 1199-B +100 −75 −75 E. faecalis +100 −87.5 −75 E. coli 2015 +100−94 −87.5 E. coli 25922 +100 −97.5 −96 P. mirabilis 37 +100 −96 −94 P.rettgeri N 1764 0.00 −94 −94 P. vulgaris 66 +100 −96 −94 Klebsiella24037 +100 −97 −94 Serratia marcescens 2702 0.00 −99 −98 Acinetobacter3109 +100 −97 −94 Ps. Aeruginosa +100 −87 −50 *% Change in Potency at pH5.5 = 100 (−MIC pH 7/MIC pH 5.5 × 100)

BIOLOGICAL EXAMPLE 3

Acute Toxicity

The acute intravenous toxicity of RS-(±)- and S-(−)- forms ofNadifloxacin in mice is shown in Table 3 below:

TABLE 3 COMPOUND LD₅₀ (mg/kg) RS-(±)-Nadifloxacin 311S-(−)-Nadifloxacin >400* *LD₅₀ 400 mg/kg i.e. no mortality observed at adose of 400 mg/kg.

BIOLOGICAL EXAMPLE 4

Hepatotoxicity Differential between S-(−)-Nadifloxacin and Trovafloxacin

Human Liver Cell Line Cytotoxicity Assay

The procedure involved cultivation of cells of human liver cell-lineHep-G2 in DMEM medium containing 5% foetal bovine and exposure tovarious concentrations of trovafloxacin and S-(−)-Nadifloxacin for 3hours. The drug containing medium was then replaced with a fresh mediumand cells were incubated in 5% CO₂ atmosphere at 37° C. for 4 days.Almar blue dye which is an indicative of active respiration was thenadded to individual sample to access the toxicity of the drugs. Thehepatotoxic potential of a drug is expressed in terms of Minimum ToxicDose (MTD) which is defined as minimum concentration of a drug whichbrings about inhibition of colour change from blue to pink.

Using above test method, S-(−)-Nadifloxacin was found to be toleratedwell by Rep-G-2 Cells at dosages 4 times higher than trovafloxacin.

BIOLOGICAL EXAMPLE 5

Bioavailability

The blood levels of RS-(±)-Nadifloxacin and S-(−)-Nadifloxacinadministered orally to Swiss mice at a dose of 30 mg/kg are shown inTable 3 with respect to the AUC (μg/ml.hr), monitored from 15 minutes to4 hours.

TABLE 4 COMPOUNDS AUC ((μg/ml · hr) RS-(±)-Nadifloxacin 16.9S-(−)-Nadifloxacin 33.58 S-(−)-Nadifloxacin has increased oralbioavailability compared to racemic Nadifloxacin.

BIOLOGICAL EXAMPLE 6

Effect of NorA Efflux Pump on Fluoroquinolone Potency

According to NCCLS protocols, comparative MICs were determined for S.aureus strain bearing Nor A efflux pump and a corresponding patentstrain devoid of efflux pump. Using this set of MIC values, % loss inpotency due to efflux was calculated for each of the fluoroquinolone inTable 5

TABLE 5 % loss in potency for S. aureus bearing Fluoroquinolone Effluxpump S-(−)-Nadifloxacin 0 Norfloxacin 97 Ciprofloxacin 94 Levofloxacin75 Gemifloxacin 75 Clinafloxacin 87.5 Gatifloxacin 75

BIOLOGICAL EXAMPLE 7

Fold Elevation in ED₅₀ Dose of Fluoroquinolones

For S. aureus with Nor A Efflux Pump

In mouse model of infection caused by S. aureus with and without effluxpump, 50% protective dosages were determined for Ciprofloxacin,Ofloxacin, Levofloxacin and S-(−)-Nadifloxacin. From theseexperimentally determined values, fold increase in 50% protective dosewas calculated and is shown in Table 6

TABLE 6 Fluoroquinolone Fold Increase Ciprofloxacin >10 Ofloxacin 10Levofloxacin 8-10 S-(−)-Nadifloxacin 0

TEST EXAMPLE 1

Equilibrium Moisture Content Determination ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H1-benzo[i,j]quinolizine-2-carboxylic acid anhydrate andS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.2 hydrate.

Silica (anhydrous) and three saturated solutions of electrolytesprepared by dissolving the respective salts in water were eachintroduced into different desiccators to control the inner relativehumidity to a specific value as represented in the following Table 7.Then, the equilibrium moisture contents ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid anhydrate andS-(−)-9-fluoro-carboxylic acid 0.2 hydrate prepared in Examples 1 and 2respectively were determined at several relative humidities.

TABLE 7 Powder/Saturated salt solutions inside the dessicator. Relativehumidity Powder/Salt Solution (%) at 27° C. Silica 20% Ammonium Nitrate58% Sodium Chloride 75% Potassium Nitrate 95%

Specifically, 1 g of the sample was spread on a pre-weighed petridishand the total weight was accurately measured, then both the samples wereplaced in each desiccator of Table 8 The dessicators were allowed tostand at normal temperature for at least 3 days and the weight wasmeasured agingly over this period. The weight changes were tabulated .At the end of 3 days all samples were taken to be weighed. The moisturecontent of each sample was determined by Karl Fischer analysis.Equilibrium moisture content at each relative humidity is represented inFIG. 1 (anhydrate) and FIG. 2 (0.2 hydrate). FIG. 2 shows that themoisture content of the 0.2 hydrate is maintained around 1% for thewhole humidity range tested (20% to 95%). FIG. 1 shows that the moisturecontent of the anhydrate is maintained around 0.1% at the relativehumidity 20% to 58%. At humidities of about 75% the anhydrate showsweight change and reaches a new equilibrium which is maintained around1% for the relative humidity range 75% to 95% and corresponds to the 0.2hydrate (by Karl Fisher measurements and X-ray diffraction analysis).The 0.2 hydrate displays superior stability since it keeps a constantequilibrium moisture content regardless of relative humidity change.

TEST EXAMPLE 2

X-ray Diffraction Analysis

After 300 mg each ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid anhydrate (prepared as in Example 1)andS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.2 hydrate (prepared as in Example2)were thinly spread on the sample holder X-ray diffraction analyses (40kv×40 mA Rigaku D/max 2200) were performed under the conditions listedbelow:

-   scan speed (2θ) 5°/min-   sampling time 7 min-   scan mode: continous-   2θ/θ reflection-   Cu target (No filter)

Results of the X-ray diffraction analysis on anhydrate and 0.2 hydratewere as depicted in FIGS. 3 and 4 respectively. From these spectra itcan be verified that their crystal forms differ from each other.

TEST EXAMPLE 3

Thermal Analysis of theS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid anhydrate (prepared as in Example 1)andS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.2 hydrate anhydride (prepared as inExample2).

For the Differential Scanning Calorimetry, PERKIN ELMER DSC 7 system wasused. 3 mg of the sample was weighed into the aluminium pan, which wasthen press sealed with an aluminium lid. After three tiny needle holeswere made on the lid the sample was tested by heating from (15° C.) to(300° C.) at a rate of 20° C./min. As can be seen from the FIG. 8 thereis an endothermic peak which begins at around 150° C., and an exothermicpeak due to thermal decomposition at around 240° C. to 264° C. Incontrast the anhydrate shows only an exothermic peak at around 245° C.to 268° C. without any endothermic peak.

TEST EXAMPLE 4

Chemical Stability Under Heating

The chemical stability ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid anhydrate (prepared as in Example 1)andS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.2 hydrate (prepared as in Example2),S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid 0.5 hydrate (prepared as in Example3) andS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.75 hydrate (prepared as in Example4) were compared in order to determine the effect on chemical stabilityof the extent of hydration.

The anhydride and hydrates were each introduced into a glass vial andmaintained at 70° C. Thus the thermal decomposition with elapsed timewas analysed by HPLC and the results thus obtained are described inTable 8

TABLE 8 Thermal stability with elapsed time at 70° C. (Unit %) Time(week) Sample Initial 1 2 Anhydrate 98.9 98.7 98.6 0.2 hydrate 98.7 98.098.1 0.5 hydrate 98.1 97.0 96.1 0.75 hydrate 98.3 97.1 97.2

As can be seen the 0.2 hydrates shows the same degree of chemicalstability as the anhydrate, whilst the 0.5 hydrate and 0.75 hydratedecompose with time.

TEST EXAMPLE 5

Water Solubility ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid anhydrate (prepared as in Example 1),S-(−)-9-Fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid 0.2 hydrate (prepared as in Example2),S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid sodium salt monohydrate (prepared asin Example 5),S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid arginine salt 0.75 hydrate (preparedas in Example 8),8-{4[2(R)-amino-3-phenylpropionyloxy]piperidin-1-yl}-9-fluoro-5(S)-methyl-6,7dihydro-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid acetate(prepared as in Example 30).

Water solubilities of the compounds listed above were measured. Themeasurement results are listed in Table 9

TABLE 9 Water Solubility at 27° C. Sample Distilled Water (pH 6.8)Anhydrate   0.19 mg/ml 0.2 hydrate   0.24 mg/ml Na Salt •H₂O >1000 mg/mlArginine Salt •0.25 H₂O    75 mg/ml Arginine Salt •0.75 H₂O    94 mg/mlD-Ala-S-(Nadifloxacin) •AcOH  >250 mg/ml

As can be seen from the above results, the salt shows superior watersolubility.

The following examples relate toS-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid, its salts, prodrugs, derivatives andhydrates thereof of the Formula I, wherein the percentages indicated inthe examples for the salts, prodrugs, derivatives and hydrate of thecompounds of the invention are calculated on the basis ofS-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid.

COMPOSITION EXAMPLE 1

Tablet composition Ingredient % w/w 1.S-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-   10-905-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylicacid or an optically pure compound of the invention of formula I. 2.Cyclodextrin & derivatives   5-40 3. Sodium citrate 0.1-5 4.Microcrystalline cellulose   1-50 5. Polyvinyl pyrrolidone 0.1-9 6.Cross carmellose sodium 0.1-5 7. Starch   2-30 8. Lactose   2-40 9.Magnesium stearate 0.1-5 10. Talc purified 0.1-5 11. Hydroxypropylmethyl cellulose 0.1-6 12. Polyethylene glycol 400 0.1-2 13. TitaniumDioxide 0.1-2

The active ingredientS-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid or an optically pure compound of theinvention of formula I is mixed with cyclodextrin and its derivatives,sodium citrate, microcrystalline cellulose, corn starch and lactose. Wetgranulate with polyvinyl pyrrolidone. Dry the granulate. Mix with crosscarmellose sodium, magnesium sterate and talc purified. Compress thetablets. Film coat the tablets using mixture of hydroxypropylmethylcellulose, polyethylene glycol 400 and titanium dioxide in appropriatesolvent.

COMPOSITION EXAMPLE 2

Injection Composition

Ingredient % w/v 1. S-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)- Up to 105-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylicacid or an optically pure compound of the invention of formula I. 2.Sodium citrate 0-3   3. Sodium hydroxide q.s. and trometamol q.s. toadjust q.s. pH between 8.0–9.9 4. Disodium edetate 0-0.5 5. Water forinjection q.s. to 100

The active ingredientS-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid or an optically pure compound of theinvention of formula I and sodium citrate is dissolved in water forinjection. Disodium edetate is added and dissolved. pH is adjusted with1% sodim hydroxide solution and trometamol. Volume to be made. Filterthrough 0.2 micron membrane filter. Fill in vials and autoclave at 121°C. for 15 minutes.

COMPOSITION EXAMPLE 3

Injection Formulation Ingredient % w/v 1.S-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)- Up to 105-methyl-6,7-dihydro-1-oxo-1H,5H-benzo [i,j] quinolizine-2-carboxylicacid or an optically pure compound of the invention of fomula I. 2.L-arginine 0.1-10  3. Sodium citrate   0-3.5 4. Sodium hydroxide toadjust pH between 8.0-9.9 q.s. 5. Disodium edetate   0-0.5 6. Water forinjection q.s. to 100

Dissolve L-arginine in water for injection. Add and dissolveS-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j] quinolizine-2-carboxylic acid or an optically pure compound of theinvention of formula I by stirring in above solution. Add sodium citrateand dissolve. Add disodium edetate and dissolve by stirring. Check pHand adjust if necessary with 1% sodium hydroxide solution. Make upvolume with water for injection. Sterilise by filtration through 0.2 μmembranes. Fill in to containers aseptically and seal.

COMPOSITION EXAMPLE 4

Topical Composition

A typical pharmaceutical cream containing 1%S-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid or an optically pure compound of theinvention of formula I was prepared using the following composition:

Ingredient % w/v S-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-0.1-10  6,7-dihydro-1-oxo-1H,5H-benzo[i,j] quinolizine-2- carboxylicacid or an optically pure compound of the invention of formula IDiethanolamine 0.1-2   Trometamol   0-0.5 Sodium hydroxide q.s. toadjust pH between 8.0-9.9 q.s. Liquid paraffin  0-20 Microcrystallinewax  0-10 Cetomacrogol 1000 0.1-5   Propylene glycol  0-20 Disodium EDTA  0-0.5 Sodium disulphite   0-0.5 Cetostearyl alcohol 0.1-15  Purifiedwater q.s. to 100

The active ingredient isS-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid or an optically pure compound of theinvention of formula I. The remaining components are inert or auxiliary.The composition of liquid paraffin, microcrystallaine wax andcetomacrogol 1000 is prepared and added to the solution ofS-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylic acid or an optically pure compound of theinvention of formula I in a mixture of diethanolamine/ trometamol. Themixture is homogenised and to the resultant cream is added propyleneglycol, sodium bisulphite and disodium EDTA. The composition is madeupto 100% with purified water to give the final composition. The creamis stable when stored at a temperature not exceeding 35° C. The pH ofstability is between 8.0 to 9.5.

1. A process for the preparation ofS(−)-9-Fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid.0.2 H₂O which comprises the steps of: a) dissolvingS(−)-9-Fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid in a volume of organic solvent, at an elevated temperature,sufficient to allow crystallization ofS-(−)-9-Fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid 0.2H₂O upon addition of water, b) adding said water in an amountsufficient to bring about crystallization after cooling, and c) cooling,filtering and drying the separated crystals at temperatures up to 40-50°C. for 3-6 hours, in vacuo upto 50 mm of Hg to a constant weight.
 2. Aprocess for the preparation ofS(−)-9-fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid 0.5 hydrate which comprises the steps of: a) dissolvingS-(−)-9-fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid in a volume of organic solvent at reflux temperature sufficient toallow crystallization of S(−)-9-fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid 0.5 hydrate upon addition of water, b) adding an amount of water atambient temperature, sufficient to bring about crystallization aftercooling and c) cooling, filtering and drying the separated crystals attemperatures up to <40° C. for 3-6 hours.
 3. A process for thepreparation of S(−)-9-fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid 0.75 hydrate which comprises the steps of: a) suspendingS-(−)-9-fluoro-8(4-hydroxypiperidin-1-yl)-5-methyl-6,7--dihydro-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid in water, b) formulating into a slurry by stirring with an organicsolvent, at 5° C. for 1-2 hours, and c) filtering and drying the productat temperatures <40° C. for 3-6 hours.
 4. A compound selected from thegroup consisting ofS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid, arginine salt;S-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid, arginine salt 0.25H₂O; andS-(−)-9-fluoro-6,7-dihydro-8-(4-hydroxypiperidin-1-yl)-5-methyl-1-oxo-1H,5H-benzo[i,j]quinolizine-2-carboxylicacid, arginine salt 0.75H₂O.
 5. A solid composition comprising acompound of claim 4 and a pharmaceutically acceptable solid carrier,diluent or excipient.
 6. A composition according to claim 5 in a formselected from the group consisting of a tablet, coated tablet, capsule,patch, powder, pill, dispersion, cachet and hard gelatin capsule.
 7. Amethod for treating a bacterial infection, mycobacterial infection, ornosocomial pathogen infection in a human or animal, which comprisesadministering to the human or animal in need thereof an amount of acompound according to claim 4 sufficient to eradicate said infection. 8.A method for treating a bacterial infection, mycobacterial infection, ornosocomial pathogen infection in a human or animal, which comprisesadministering to the human or animal in need thereof an amount of acomposition according to claim 5 sufficient to eradicate said infection.9. The method according to claim 7 wherein the bacterial infection iscaused by a resistant Gram-positive organism or a Gram-negativeorganism.